CROSS-REFERENCE TO RELATED APPLICATIONS The present application claims the benefit of U.S. Provisional Application No,

62/366,539, filed on My 25, 2016, and U.S. Provisional Application No. 62/527,415, filed June 30, 2017, each of which is incorporated herein by reference in its entirety.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has been submitted electronically in ASCII format and is hereby incorporated by reference in its entirety. Said ASCII copy, created on July 25, 2017, is named 12444 J)683-00304JSL.txt and is 817,847 bytes in size.

TECHNICAL FIELD

This disclosure relates to materials and methods for biosynthesizing one or more compounds having an odd number of carbon atoms, for example, five to nineteen carbon atoms. This disclosure also relates to materials and methods for biosynihesizing one or more difunctional products having an aliphatic carbon chain backbone having an odd number of carbon atoms between five and nineteen inclusive and. having two terminal functional groups selected from carboxyl, formyl, amine, and hydroxy! or salts or derivatives thereof (hereafter "C5-C19 building blocks") from propanedioyl-CoA or propanedioyl- [acp] and optionally acety!-CoA using one or more polypeptides having the activity of one or more enzymes such as methy [transferases, [i~keioacyl~[acp] synthases, β-ketothiolases, dehydrogenases; reductases, hydrolases, thioesterases, esterases, CoA-transferases, reversible CoA-Ugases, and aminotransferases or using recombinant host cel ls expressing one or more nucleic acids encoding such enzymes in genetically modified hosts. The disclosure provides biochemical pathways in which a methyl ester shield is added to propanedioyl-CoA or propanedioyl- [acp] before undergoing one or more cycles of carbon chain elongation, wherein the methyl ester shield is maintained for at least one further enzymatic step following the one or more cycles of carbon chain elongation, BACKGROUND

Nylons are syntheti polyamides which are sometimes synthesized by the condensation polymerisation of a diamine with a dicarboxylic acid. Similarly, Nylons may be produced by the condensation polymerisation of lactams. For example, a ubiquitous Nylon is Nylon 6,6, which is produced by reaction of hexamethy!enediamine (HMD) and adipic acid. Nylon 6 is produced by a ring opening polymerisation of caprolactam (Anton & Baird, Polyamides Fibers, Encyclopedia of Polymer Science and Technology, 2001).

Given the lack of economically cost competitive petrochemical monomer feedstocks, biotechnology offers an alternative approach via biocaialysis. Biocaialysis is the use of biological catalysts, such as enzymes, to perform biochemical transformations of organic compounds. Both bioderived feedstocks and petrochemical feedstocks are viable starting materials for the biocaialysis processes.

However, no wild-type prokaryote or eukaryote naturally overproduces or excretes difutictionai products having an odd number of carbon chain atoms, such as, for example, C _¾ - Ct9 building blocks, to the extracellular environment. Nevertheless, the metabolism of the C? dicarboxylic acid (heptanedioie acid) has been reported.

The C _? dicarboxylic acid, heptanedioie acid, is converted efficiently as a carbon source by a number of bacteria and yeasts via β-oxidation into central metabolites, β- oxidation of CoBnzyme A (Co A) activated heptanedioate to CoA-activated 3- oxoheptanedioate facilitates further catabolism via, for example, pathways associated with aromatic substrate degradation. The catabolism of 3 -oxopimeloyi-Co A to aeetyi-CoA and gSutaryl-CoA by several bacteria has been characterized comprehensively (Harwood and Parales, Annual Review of Microbiology, 1996, 50, 553 - 590).

The optimally principle states that microorganisms regulate their biochemical networks to support maximum biomass growth. Beyond the need to express heterologous pathways in a host organism, directing carbon flux towards C5-C19 building blocks (i.e., Cj, C-7, C9, Ct>., Ci3, C35, Cj7, or Ci building blocks) that serve as carbon sources rather than to biomass growth constituents, contradicts the optimaiit principle. For example, transferring the 1 -butanol pathway from Clostridium species into other production strains has often fallen short by an order of magnitude compared to the production performance of native producers (Shen el al, , Appl Environ. Microbiol. , 2011 , 77(9), 2905 - 2915). The synthesis of an aliphatic carbon backbone precursor having an odd number of carbon atoms, for example, between five and nineteen carbons, is a key consideration in synthesizing difunctional products having an odd number of carbon atoms (i.e., C5-C39 building blocks) prior to forming terminal functional groups, such as carboxyl, formyl, amine, or hydroxy! groups, on the Cs-C^ aliphatic backbone.

SUMMARY

Accordingly, against this background, it is clear that there is a need for methods for producing products having two terminal functional groups and an odd number of carbon chain atoms, or salts or derivatives thereof, wherein the methods are biocaialyst-based . For example, there is a need for methods for producing difunctional products having an odd number of carbon, atoms between five and nineteen (i.e., 5, 7, 9, 1 ί , 13, 15, 17, or 19 carbon atoms) and two terminal functional groups (hereafter C- C j o building blocks). Described herein are methods and genetically modified hosts that allow for more efficient use of five to nineteen carbon aliphatic backbone precursors and the production of C ₅ "C j budding blocks, for example, by use of recombinant host in a hioH deficient background.

This document is based at least in part on the discovery that it is possible to construct biochemical pathways for producing a five to nineteen carbon chain aliphatic backbone precursor (i.e., an aliphatic backbone with 5, 7, 9, 11, 13, 15, 17, or 19 carbon atoms), in which one or two functional groups, i.e., carboxyl, formyl, amine, or hydroxy!, can be formed, leading to the synthesis of one or more products having two terminal functional groups, or salts or derivatives thereof (C5-C19 building blocks). These may be, for example, dicarboxy!ic acids, carboxylase semialdehydes, aminocarboxy!ates, hydroxycarboxylat.es, diamines, or diols. The compound may exist in any of its neutral or ionized forms, including any salt, forms thereof. It is understood by those skilled in the art thai the specific form will depend on pH.

For compounds containing carboxyiic acid groups such as organic monoacids, hydroxyacids, aminoaeids, and dicarboxylic acids, these compounds may be formed or converted to their ionic salt form when an acidic proton present in the parent compound either is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion; or coordinates with an organic base. Acceptable organic bases include ethanolamine, diethanolamine, triethanolamine, tromethaniine, -methyiglucamine. and the like. Acceptable inorganic bases include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, sodium hydroxide, and the like. The salt can be isolated as is from the system as the salt or converted to the free acid by reducing the pH to below the pKa through addition of acid or treatment with an acidic ion exchange resin.

For compounds containing amine groups such as but not limited to organic amines, aminoacids and diamines, these compounds may be formed or converted to their ionic salt form by addition of an acidic proton to the amine to form the ammonium salt, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; or formed with organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropioriic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maieic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethancsulfonic acid, 1 ,2-ethanedisu!fonic acid, 2-hydroxyethanesuifbnic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-methylbicycio-[2.2.2]oct-2 ^» ene-l-carboxylic acid, glucoheptonic acid, 4,4'~methylenebis-(3 -hydroxy-2-ene- 1 -carboxyiic acid), 3- pheny [propionic acid, trimethylacetic acid, tertiary butyl acetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphtnoic acid, salicylic acid, stearic acid or muconic acid. Acceptable inorganic bases are known in the art and include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, sodium hydroxide, and the like. The salt can be isolated as is from the system as a salt or converted to the free amine by raising the pH to above the plvb through addition of base or treatment with a basic ion exchange resin.

For compounds containing both amine groups and carboxyiic acid groups such as but not limited to aminocarboxylates, these compounds may be formed or converted to their ionic salt form by either 1) acid addition salts, formed with inorganic, acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; or formed with organic acids such as acetic acid, propionic acid, hexanoic acid, eycloperttanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maieic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3~(4- hydroxybenzoyiybenzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1 ,2-ethanedisuifonic acid, 2-hydroxyethanesulfonie acid, benzenesulfonic acid, 2-naphthajenesulfonic acid, 4-methylbicyclo-[2.2.2]oct~2-ene-l-carboxylic acid, glucoheptonic acid, 4,4'-methylenebis-(3-hydroxy-2-ene-l-carboxylic acid), 3- phenylpropionic acid, trimethylacetic acid, tertiary bulylacetic acid, lauryl sulfuri acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, m conic acid Accepiable inorganic bases include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, sodium hydroxide, and the like, or 2} when an acidic proton present in the parent compound either is replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or an aluminum ion; or coordinates with an organic base. Accepiable organic bases are known in the art and include ethanolamine, diethanolamine, tri ethanolamine, tromethamine, M-methylglucamine, and the like. Acceptable inorganic bases are known in the art and include aluminum hydroxide, calcium hydroxide, potassium hydroxide, sodium carbonate, sodium hydroxide, and the like. The salt can be isolated as is from the system or converted to the free acid by reducing the pH to below the pKa through addition of acid or treatment with an acidic ion exchange resin.

The pathways, metabolic engineering strategies, and cultivation strategies described herein rely on fatty acid elongation and synthesis e zymes or homologs accepting methyl- ester shielded dicarboxylic acids as substrates.

In the face of the optimality principle, it surprisingly has been discovered that appropriate non-natural pathways, feedstocks, host microorganisms, attenuation strategies to the host's biochemical network and cultivation strategies may be combined to efficiently produce one or more diiunctional products having an odd number of carbon atoms, such as, for example, CJ-CJ building blocks.

Each of said one or more cycles of carbon chain elongation to produce the aliphatic carbon chain backbone having an odd number of carbon atoms, such as, for example, an aliphatic carbon chain backbone five to nineteen carbons in length, comprises using (i) a β~ ketoaeyl~[acp] synthase or a β-ketothiolase, (it) a 3-oxo cyI-f cpJ reductase, an acetoacetyl- CoA reductase, a 3 -hydroxyacyl-CoA dehydrogenase or a 3-hydroxybutyryl-CoA dehydrogenase, (iii) an enoyl-CoA hydr tase or a 3-hydroxyacyl-[ cp] dehydratase, and (iv) ati enoyl-f cp] reductase or a trans-2-enoyI-CoA reductase to produce said aliphatic carbon chain backbone from propanedioyl-[acp] methyl ester or from propanedioyhCoA methyl ester. A S~adenosyl-L-m t ionine (SAM) -dependent m thyiiramfer se may convert propanedioyl-CoA to propanedioyl-CoA methyl ester or convert propanedioyl-[acp] to propanedioyl~[acp] methyl ester before said one or more cycles of methyl-ester shielded carbon chain elongation.

n some embodiments, a €2*1-3 aliphatic backbone containing (2«+3) carbon atoms, wherein n is an integer greater than or equal to one, is enzymatically synthesized from (i) acetyl-CoA and propanedioyi-CoA via n cycles of methyl ester shielded carbon chain elongation or (ii) propanedioyl-jacp] via n cycles of methyl ester shielded carbon chain elongation. In some embodiments, a G2n+3 aliphatic backbone may be formed from propanedioyi-[aep] or propanedioyl -Co A, and optionally acetyi-CoA, via one or more cycles of carbon chain elongation using polypeptides having the activity of one or more either NADH or NADPH dependent enzymes.

For example, €5. C9, Cn, C13, € ₁₅ , (h?, or C;o aliphatic backbones may be enzymatically synthesized from (i) acetyl-CoA and propanedioyl-CoA via 1. 2, 3, 4, 5, 6, 7, or 8 cycles of methyl ester shielded carbon chain elongation, respectively, or (ii) propanedioyl- [ac-p] via 1 , 2, 3, 4, 5, 6, 7, or 8 cycles of methyl ester shielded carbon chain elongation, respectively, €5, C7, C9, Cn, Cn, C _\ $, Cj _? , or Cj9 aliphatic backbones disclosed herein include; i) C5: pentanedioyl-[acp] methyl ester or pentanedioyl-CoA methyl ester, ii) C ₇ : heptanedioyl-[acp] methyl ester or heptanedioyi-CoA meihyl ester, iii) (¾: nonanedioyh[acp] methyl ester or nonanedioyl-CoA methyl ester, iv) Cj < : undecanedioyi-[aep] methyl ester or undecanedioyl-CoA meihyl ester, vj C13: tridecanedioyi-[acp] methyl ester or tridecanedioyi- CoA methyl ester, vi) s: pentadecanedioyl-[aep] methyl ester or pentadecanedioyi-CoA methyl ester, vii) Ci?: heptadecanedioyl-[acp] methyl ester or heptadecanedioyl-CoA methyl ester, or viii) C 1 : nonadecanedioyl-jacp] methyl ester or nonadecanedioyl-CoA meihyl ester.

In some embodiments, a Can-t-s aliphatic backbone is converted to one or more C?. _R +3 building blocks. As used herein, a "¾„ ₊ ¾ building block" is a difunctional compound having (2«+3) carbon atoms, wherein n is an integer greater than or equal to one, and two terminal functional groups selected from carboxyl, forrnyl, amine, and hydroxyl groups. Difunctional compounds having an odd number of carbon atoms described herein include dicarboxylie acids, carboxylate semialdehydes, aminocarboxylates, hydroxyearboxylates, diamines, and diols, In some embodiments, a terminal carboxyl group can be enzymatically formed using one or more polypeptides having the activity of one or more esterase, thioesterase, aldehyde dehydrogenase, 7~oxoheptanoate dehydrogenase, 6-oxohexanoate dehydrogenase, reversible Co A ligase (e.g., reversible succin l-CoA ligase), or CoA-transferase (e.g., glutaconaie CoA- transferase),

in some embodiments, a terminal amine group can be enzymatically formed using a polypeptide having the activity of an aminotransferase or a deacetylase.

In some embodiments, a terminal hydroxy! group can be enzymatically formed using a polypeptide having the activity of a 4~hydroxybutyrate dehydrogenase, a 5- hydroxypentanoate dehydrogenase, a 6-hydroxyhexano te dehydrogenase, or an alcohol dehydrogenas .

in one aspect, this document features a method for biosynthesizing a difunctional product having an aliphatic carbon backbone with an odd number of carbon atoms (e.g., between five and nineteen carbon atoms) and two terminal functional groups selected from carboxyl formyl, amine, and hydroxy! groups, or salts or derivatives thereof.

The difunctional product having an odd number of carbon atoms may be selected from dicarboxylic acids, carboxylase semialdehydes, aminocarboxylaies, hyd oxyearboxylates, diamines, and diols. For example, the difunctional product may be selected from pentanedioic acid, 5-oxopentanoate, 5-arninopentanoate, 5 -hydroxypentanoate, pentane~l,5~ diamine, and L5~pentanediol: heptanedioic acid, 7-oxoheptanoate, 7-aminoheptanoate, 7- hydroxyheptanoate, heptane- 1,7-diamine, and 1 ,7-heptanediol; nonanedioic acid, 9- oxononanoate, 9-aminononanoate, 9-hydroxynonanoate, nonane- 1 ,9~diamine, and 1,9- nonanedioi; undecanedioic acid, l l-oxoundecanoate, 1 ϊ-aminoundecanoate, 11- hydroxyundecanoate, undecane- 1 , 1 1 -diamine, and 1 ,1 i-undecanediol; tridecanedioic acid, 13- oxotridecanoate, 13-aminotridecanoate, 13-hydroxytridecanoate, tridecane- 1 , 13-diamine, and 1,13-tridecanediol; pentadecanedioic acid, 15-oxopentadecanoate, 15-a inopentadeeanoate, 15-hydroxypentadecanoate, pentadecaiie- 1 , 15-diamine, and 1 ,15-pentadecanediol; heptadecanedioic acid. ί 7-oxoheptadecanoate, 17-aminoheptadecanoate, 17- hydroxyheptadecanoate, heptadecane- 1 , 17-cHamine, and 1 , 17-heptadecanediol; nonadecanedioic acid, 19-oxononadecanoate, 19-aminononadecanoate, 19- hydroxynonadecanoate, nonadeca.ne-1 ,1 9-diamine, and 1 ,19-nonadeeanedioi. In some embodiments, the method includes (a) enzymatically synthesizing an aliphatic carbon chain backbone having an odd number of carbon atoms from (i) acetyl-CoA and propanedioyl-CoA via one or more cycles of methyl ester shielded carbon chain elongation or (it) propanedioyl-[acp] via one or more cycles of methyl ester shielded carbon chain elongation, (b) enzymatically forming a first terminal functional group selected from carboxyi, amine, formyl, and hydroxy! groups in said backbone while maintaining said methyl ester shield for at. least one farther enzymatic step, and (c) enzymatically forming a second terminal functional group selected from carboxyi, amine, formyl, and hydroxy! groups in said backbone, thereby forming said difunctional product, in some embodiments, the number of carbon atoms in the backbone may be any of five, seven, nine, eleven, thirteen, fifteen, seventeen, or nineteen carbon atoms.

Following one, two, three, four, five, six, seven, or eight cycles of methyl ester shielded carbon chain elongation, the following aliphatic backbones may be formed: pentanedioyl-jaep] methyl ester or pentanedioyi-CoA methyl ester; heptanedioyI-[acp] methyl ester or heptanedioyl-CoA methyl ester; nonanedioyl-[acp] methyl ester or nonanedioyl-CoA methyl ester; undecanedioyl-[acp] methyl ester or undeeanedioyl- CoA methyl ester; tridecanedioyl-[acp] methyl ester or trideeanedioyl~CoA methyl ester; pentadecanedio l-jaep] methyl ester or pentadecanedioyi-CoA methyl ester; heptadecanedioyl-jaepj methyl ester or heptadecanedioyl-CoA methyl ester; or nonadeeanedioyl- [acp j methyl ester or nonadecanedioyl-CoA methyl ester.

Each of the aforementioned aliphatic backbones can be converted to the respective central precursor, namely pentanedioyl-jacp] or pentanedioyi-CoA, heptanedioy 1- [acp] or heptanedioyl-CoA, nonanedioyl-[aep] or nonanedioyl-CoA, undeeanedioyl- [acp] or undecanedioyl-CoA, tridecanedioyl~[acp] or tridecanedioyl-CoA, pentadecanedioyI-[acp] or pentadecanedioyl-CoA, heptadecanedioyl-[aep] or heptadecanedioyl-CoA, nonadecanedioyl- [aep] or nonadeeanedioyl-CoA by a polypeptide having the activity of [aepjmethyl ester esterase classified, for example, under EG 3.1.1.85, such as the gene product of hioH, from, for example Escherichia coli (see UniProiKB Accession No. P13001 (SEQ ID NO: 139). However, some analyses indicate that a thioesterase may bypass the activity of BioH and instead produce a monomethyi carboxylate and holo~[ACP] or hoio-CoA (see, e.g., FIG. 36). Since BioH may show low activity towards monomethyi carboxylate, this process may lead to a decrease in carboxylate production. In some embodiments, the method comprises synthesizing an aliphatic carbon chain backbone via one or more cycles of methyl ester shielded carbon chain elongation in a bioH deficient background. In some embodiments, the method comprises a step of do nregulating the activity of bioH. For example, in some embodiments, the method is performed in a recombinant host comprising a deletion in bioH. In some embodiments, the recombinant host does not express BioH.

In any of these embodiments, after the one or more cycles of carbon chain elongation, the methyl ester shield may be maintained for at least one further enzymatic step. In some embodiments, the at least one further enzymatic step comprises the enzymatic conversion of methyl ester intermediates to the respective monomethvl carboxylate.

Therefore, in some embodiments, the resulting pentanedioyl-[acp] methyl ester or pentanedioylCoA methyl ester, heptanedioyl-[acp] methyl ester or heptanedioyl-CoA methyl ester, nonanedioyl-[acp] methyl ester or nonanedioyl-CoA methyl ester, undecanedioyl-[acp] methyl ester or undeeanedio l-CoA methyl ester, tridecanedioyl- acp] methyl ester oi ^¬ iridecanedioyl-C A methyl ester, pentadecanedioyl~[acp] methyl ester or pentadecanedioyl- CoA methyl ester, heptadeeanedioyhj acp] methyl ester or heptadecanedioyl-CoA methyl ester, or nonadecanedio l- acp] methyl ester or nonadeeanedioyi-CoA methyl ester can be further converted to the respective monometbyi carboxylate by a polypeptide having the activity of thioeslerase classified, for example, under EC 3.1.1 ,2, EC 3, 1.1 ,5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2,21. or EC 3.1.2.27,

In one aspect, pentanedioy!~[aepj methyl ester or pentanedioyl-CoA methyl ester can be converted to monomefhyl pentanedioate; hepianedioyl-[aep] methyl ester or heptanedioyl- CoA methyl ester to monomethyl heptanedioate; nonanedioyl-[acp] methyl ester or nonanedioyl-CoA methyl ester to monomethyl nonanedioate; utidecanedioyl-[acp] methyl ester or undecanedioyl-CoA methyl ester to monomethyl undecanedloate; iridecanedioyl- [acp] methyl ester or tridecanedioybCoA methyl ester to monomethyl tridecanedioate; pentadecanedioy[-[aep] methyl ester or peniadecanedioy!-CoA methyl ester to monomethyl pentadecanedioate; heptadecanedioyl-[aep] methyl ester or heptadeeanedioybCoA methyl ester to monomethyl heptadecanedioate; and nonadecanedioy]-[acp] methyl ester or nonadeeanedioyi-CoA methyl ester to monomethyl nonadecanedioate, In some embodiments, the at least one further enzymatic step comprises the enzymatic conversion of a monomethyl carboxylate to a rnonomethvi carboxylate semialdehyde. In some embodiments, the at least one further enzymatic step comprises the enzymatic conversion of monomethyl perrtanedioate to methyl 5-oxopentanoate; the enzymatic conversion of monomethyl heptanedioate to methyl 7-oxoheptanoate; the enzymatic conversion of monomethyl nonanedioate to methyl 9-oxononanoate; the enzymatic conversion of monomethyl nndecanedioate to methyl 11 -oxoundecanoate; the enzymatic conversion of monomethyl tridecanedioate to methyl 13-oxotridecanoate; the enzymatic conversion of monomethyl pentadeeanedioate to methyl 15-oxopentadecanoate; the enzymatic conversion of monomethyl heptadecanedioate to methyl 17-oxoheptadecanoate; or th enzymatic conversion of monomethyl nonadecanedioate to methyl ί 9-oxononadecanoate.

in some embodiments, the at least one further enzymatic step comprises the enzymatic conversion of a monomethyl carboxylate semialdehyde to a monomethyl aminocarboxylate. in some embodiments, the at least one further enzymatic step comprises the enzymatic conversion of methyl 5-oxopentanoate to monomethyl 5-aminopentanoate; methyl 7- oxoheptanoate to monomethyl 7-aminoheptanoate; methyl 9-oxononanoate to monomethyl 9~ aminononanoate; methyl 11 -oxoundecanoate to rnonomethvi l l -aminoundecanoate; methyl 13-oxotridecanoate to monomethyl 13-aminotridecanoate; methyl 15-oxopentadecanoate to monomethyl 15-aminopentadecanoate; methyl i 7-oxoheptadecanoate to monomethyl 17- aminoheptadecanoate; or methyl 19-oxononadecanoate to monomethyl 19- aminononadecanoate.

In some embodiments, the at least one further enzymatic step compri es the enzymatic conversion of the aliphatic carbon chain backbone to a monomethyl carboxylate semialdehyde. In some embodiments, the following enzymatic conversions may occur: penta.nedioyl~[aep] methyl ester or pentanedioyl-CoA methyl ester can be converted to methyl 5-oxopentanoate; heptanedioyl-[acp] methyl ester or heptanedioyl-CoA methyl ester to methyl 7-oxoheptanoate; nonanedioybjacp] methyl ester or nonanedioyl-CoA methyl ester to methyl 9-oxononanoate; undecanedioyl-[acp] methyl ester or undecanedioyi-CoA methyl ester to methyl 11 -oxoundecanoate; tridecanedioyl-[acp] methyl ester or trideeanedioyi-CoA methyl ester to methyl 13-oxotridecanoate; pentadeeanedioyl-[acp] methyl ester or pentadecanedioyl- CoA methyl ester to methyl 15-oxopentadecanoate; heptadecanedioyl-[acp] methyl ester or hepiadecanedioyl-CoA methyl ester to methyl 17-oxoheptadecanoate; and nonadecanedioyl- acp] methyl ester or nonadecanedioyl-CoA methyl ester to methyl 19~oxononadeeanoate, in some embodiments, a mcmomefhyl earboxylate can be converted to the respective dicarboxylie acid using an esterase, in an embodiment, the following enzymatic conversions may occur: monornethyl pentanedioate to pentanedioic acid; monornethyl heptanedioate to heptanedioic acid; monornethyl nonanedioate to nonanedioic acid; monornethyl undecanedioate to undeeanedioic acid; monornethyl tridecanedioate to tridecanedioic acid; monomethylpentadecanoate to pentadecanedioic acid; monornethyl heptadecanedioate to heptadecanedioic acid, and monornethyl nonadecanedioate to nonadecanedioic acid.

In some embodiments, the monornethyl earboxylate semialdehyde can be converted to a earboxylate semialdehyde using a polypeptide having the activity of an esterase. In some embodiments, the following enzymatic conversions may occur: methyl 5-oxopentanoate to 5- oxopentanoate; methyl 7-oxoheptanoate to 7-oxoheptanoate; methyl 9-oxononanoate to 9- oxononanoate; methyl 11 -oxoimdecanoaie to 11-oxoundecanoate; methyl 13-oxotridecanoate to 13-oxotridecanoate; methyl ί 5-ox.opentadecanoate to 15-oxopeiitadecanoate; methyl 17- oxohepladeeanoate to 17-oxoheptadecanoate and methyl 19~oxononadecanoate to 19- oxononadecanoate.

In some embodiments, a monornethyl aminocarboxylate can be converted to an aminocarboxylate using a polypeptide having the activity of an esterase, in some embodiments, the following enzymatic conversions may occur: monornethyl aminopentanoate to 5 -aminopentanoate; monornethyl aminoheptanoate to 7-aminoheptanoate; monornethyl aminononanoate to 9~aminononanoate; monornethyl aminoundecanoate to 11 - aminoundecanoate; monornethyl aminotridecanoate to 13-aniinotridecanoate; monornethyl aminopentadecanoate to ί 5-aminopentadecanoate; monornethyl aminoheptadecanoate to 17- aminoheptadecanoate, or monornethyl aminononadecanoate to 19-aminononadeeanoate.

A dicarboxylie acid derived from a monornethyl earboxylate by the enzymatic activity of an esterase may itself be converted to a earboxylate semialdehyde, which may subsequently be converted in a further enzymatic step to an aminocarboxylate. In some embodiments, the following enzymatic conversions may occur: pentanedioic acid to 5- oxopentanoate to 5 -aminopentanoate; heptanedioic acid to 7-oxoheptanoate to 7- aminoheptanoate; nonanedioic acid to 9-oxononanoate to 9-aminononanoate; undecanedioie acid to 11 -oxoimdecanoate io 11. -aminoundecanoate; tridecanedioic acid to 13- oxotridecanoate to 13-ammotridecanoaie; pentadecanedioic acid to 15-oxopentadecanoaie to 15-aminopentadecaiioate; heptadecanedioic acid to 17-oxoheptadecanoate to 17- aminoheptadecanoate; or nonadeeanedioic acid to 19-oxononadecanoate to 19- aminononadecanoate.

In some embodiments, an aminocarboxylate may be converted to an acetamidocarboxylate, In some embodiments, the following enzymatic conversions may occur: 5-aminopenianoaie to N5-acetyl-5-aminopentanoate; 7-aminoheptanoate to N7-acetyl~ 7-aminoheptanoate; 9~aniinononanoaie to N9-aceiyl~9~aminononanoate; 11- aminoundecanoate to 11 -acetvS-11 -aminoundecanoate; 13-aminotridecanoate to Nl 3 -acet ^y l- 13 -aminotridecanoate; 15-aminopentadeeanoate to N15-acetyi-15~aminopentadecanoate; 17- aniinoheptadecanoate to 17-acetyl- ί 7-aminoheptadecanoate; or 19-aminononadecanoate to 19-acetyi- 19-aminononadecanoate.

in some embodiments, an acetamidocarboxylate may be converted to a diamine. For example, in some embodiments, an acetamidocarboxylate may be converted to an acetamidoaldehyde, followed by conversion to an aeetamidoamme, followed by conversion to a diamine, in some embodiments, the following enzymatic conversions may occur: N5- acetyl-5 -aminopentanoate to N5-aeetyl-5-aminopentanai to N5-aceiyl- 1 ,5-diaminopentane to pentane- 1 ,5-diamme; N7-acetyl~7~aminoheptanoate to N7-acetyl-7-aminohepianal to N7- acet l-1 ,7-diaminoheptane to heptane- 1 ,7-diamine; N9-acety!-9-aminononanoate to 9- aeetyl-9-ammononanal io N9-acetyl- 1 ,9-diarainonoane to nonane- 1 ,9-diamine; Nl l-acetyl- 11 -aminoundecanoate to Ni l -acetyl- 11 -ami noundecanal to Ni l -acetyl- l,l l~diaminoundecane to undecane- 1 ,11 -diamine; N13-acetyl-13-ammotrideeaiioate to N13-aeetyl-13- armnotridecanal to Nl 3 -acetyl-1 , 13-diaminotridecane to tridecane- 1 , 13-diamine; N15-acetyl- 15-aminopentadecanoate to N15-acetyl-15-aminopentadecanal to NI 5-acetyl-L15- diaminopentadecane to pentadecane-L15~diamine; Nl 7-acetyl- 17-aminoheptadecanoate to 17-acetyl- 17-aminoheptadecanal to Nl 7-acetyl- 1,17-diaminoheptadecane to heptadecane- 1,17-diarnine; or l 9-acetyi- 19-aminononadecanoate to 9-acetyl- 19-aminononadecanal to 19-acetyi- 1 , 19-diaminononadecane to nonadecane- 1.19-diamine.

in some embodiments, a carboxylate semialdehyde may be converted to a hydroxycarboxylate. In some embodiments, the following enzymatic conversions may occur; pentanoate semialdehdye io 5-hydroxypentanoate; 7-oxoheptanoate to 7- hydroxyheptanoate; 9-oxononanoat to 9-hydroxynonanoate; 11 -oxoundecanoate to 11 - hydroxyundecanoate; 13-oxotridecanoate to 13-hydroxytridecanoate; i 5-oxopentadecanoate to ί 5-hydroxypentadecanoate; 17-oxoheptadecanoate to 17-hydrox heptadecanoate, or 19- oxononadeeanoate to 19-hydroxynonadecanoate.

In some embodiments, a hydroxycarboxylate may be converted to a dial. In some embodiments, the following enzymatic conversions may occur; 5 -hy droxypentanoate to 1,5- penianedial; 7-hydroxyheptanoate to 1 ,7-heptanedial; 9-hydroxynonanoate to 1 ,9-nonanedial; 11. -hydroxyundecanoate to 1, 1-undecanedial; 13-hydroxytridecanoate to 1 ,13-tridecanedial; 15-hydroxypentadecanoate to 1 , 15-penladecanedial; 17-hydroxyheptadecanoate to 1, 17- heptadecanedial; or 19-hydroxynonadecanoate to 1 , 19-nonadecanedial.

In some embodiments, a dial may be converted to an aminoa!dehyde, in some embodiments, the following enzymatic conversions may occur: 1,5-pentanedial to 5- aminopentanal; 1 ,7-heptanedial to 7-aminoheptanal ; t ,9-nonanedial to 9-aminononanal; 1,11- undeeanedial to 11 -aminoundecanal; 1 ,13-tndeeanediaI to 13 -aminotridecanai; 1,15- pentadecanedial to 15-aminopentadecanal; 1 , 17-heptadecanediai to 17-aminoheptadecanal; or 1 ,19-nonadecanedial to 19-aminononadeeanaL

In some embodiments, an aminoaldehyde may be converted to a diamine. In some embodiments, the following enzymatic conversions may occur: 5-aminopentanal to pentane- 1,5-diamine; 7-aminoheptanai to heptane- 1 , 7-diamine; 9-aminononanai to nonane-1 ,9- diamine; ί 1 -aminoundecanal to undecane- 1 ,11 -diamine; 13~aminotridecanai to tridecane- 1,13 -diamine; 15-a.mmotridecanal to tridecane- 1.13 -diamine; 15-aminopentadecanal to pentadecane-l ,15-diamine; 17-aminoheptadecanal to heptadecane- 1 ,17-diamine; or 19- aminononadeeanal to nonadecane~l J 9~diamine.

In some embodiments, a hydroxycarboxylate may be converted to a bydroxyaidehyde. In some embodiments, the following enzymatic conversions may occur: 5-hydroxypentanoate to 5-hydroxypentanah 7-hydroxyheptanoate to 7-hydroxyheptanal; 9-hydroxynonanoate to 9- hydroxynonanal; 11 -hydroxyundecanoate to 11-hydroxyundecanai; 13-hydroxytndecanoate to 13-hydroxytridecanal; 1 -hydroxypentadecanoate to 15-hydroxypentadecanal; 17- hydroxyheptadecanoate to 17-hydroxyheptadeeanal; or 19-hydroxynonadecanoate to 19- hydroxvnonadecanal. In some embodiments, a hydroxyaldehyde may be converted to a diol, In some embodiments, the following enzymatic eonversions may occur: 5-hydroxypenlanai to 1,5- pentanediol; 7-bydroxyheptanal to 1,7-heptanediol; 9-hydroxynonanai to 1 ,9-nonanedioi; 11- hydroxyundeeanal to ij l-undecanedioi; 13-hydroxytridecanai to 1 ,13-tridecanediol; 15- hydroxypentadeeanal to 1 ,15-pentadeeanediol; 17-hydroxyheptadecanal to 1 ,17- heptadecanedioli or ί 9-hydroxynonadecanal to 1 , 19-nonadecanediol.

n some embodiments, a hydroxyaldehyde may be converted to a hydroxyamine. In some embodiments, the following enzymatic conversions may occur: 5-hydroxypentanal to 5- aminopentanol; 7-hydroxyheptanal to 7-aminoheptano I ; 9~hydroxynonanal to 9- aminononanol; 11 -hydroxyundeeanal to 1 1 -aminoundecanol; 13 -hydroxy trldecanal to 13- aminotridecanol 15-hydroxypentadecanai to 15-amiiiopentadecanoI; 17-hydroxyhepiadecanal to 17-a.minoheptadecanol: or 19-hydroxynonadecanal to 19-aminononadecanol.

In some embodiments, a hydroxyamine may be converted to an aminoaldehyde. In some embodiments, the following enzymatic conversions may occur: 5-amincpentanoi to 5- a ninopentanal; 7-aminoheptanol to 7-aminoheptaiial: 9-aminononanol to 9-aminononanal; l l-am oimdecanol to 11 -amino mdecanal; 13~aminotridecanol to 13-aminoiridecanal; 15- aminopentadecanol to 15-aminopentadecanal; 17-aminoheptadecanol to 17- aminoheptadecanal: or 1 -aminononadecanol to 19-aminoheptadeeanal,

In some embodiments, the method comprises (a) enzymatically synthesizing an aliphatic backbone having an odd number of carbons from (i) acetyl-CoA and propanedioyl- CoA via one or more cycles of methyl ester shielded carbon chain elongation or (ii) propanedioyl-jaep] via one or more cycles of methyl ester shielded carbon chain elongation, and maintaining the methyl ester shield for at least one further enzymatic step following the one or more cycles of carbon chain elongation, and (b) enzymatically forming two terminal functional groups selected from, carboxyl, formyl, amine, and hydroxyl groups in the backbone, thereby forming the difunctional product, and wherein the method comprises a step of downregulaiing the activity of met J, a methionine repressor protein that inhibits the initial step of adding a methyl ester shield to propanedioy!-CoA or propanedioyi-[acp]. For example, in some embodiments, the method is performed in a recombinant host comprising a deletion in met J. In some embodiments, the recombinant host does not express MetJ. In some embodiments, the aliphatic backbone having an odd number of carbon atoms is a five to nineteen carbon chain aliphatic backbone.

One advantage of performing a method according to this disclosure is that, in a bioH deficient background, the carboxy!-[acp] methyl ester or the earhoxy!-CoA methyl ester can be converted to the respective monomethyl carboxyiate, the respective monomethyi carboxyiate semialdehyde. and/or the respective monomethyl aminocarboxylate. Such methyl ester shielded building blocks (I.e., methyl ester shielded C CIQ building blocks) may be further converted to a carboxylic acid, carboxyiate semialdehyde, or aminocarboxylate by a suitable esterase with high efficiency, which may ultimately lead to higher yields of difunctional products having an odd number of carbon atoms.

A five to nineteen carbon chain aliphatic backbone according to this disclosure can be any of pentanedioy!-[acp] methyl ester or pentanedioyl-CoA methyl ester; heptanedioyl-j acp] methyl ester or heptanedioyl-CoA methyl ester: nonanedioyl-[acp] methyl ester or nonanedioyl-CoA methyl ester; undecanedioy 1- [acp] methyl ester or undecanedioyl- CoA methyl ester; tridecanedioyl- [acp] methyl ester or tridecanedioyl-CoA methyl ester: pentadecanedioyl~[acp] methyl ester or pentadecanedioyl-CoA methyl ester; heptadeea.nedioyi~[aep] methyl ester or heptadecanedioyl-CoA methyl ester or nonadecanedioyl-[acp] methyl ester or nonadecanedioyl-CoA methyl ester.

in some embodiments, a polypeptide having the activity of S-adenosyl-L-methioni (SAM)~dependent methyltransferase can convert propanedioyl-CoA to a propanedioyl-CoA methyl ester or can convert propanedioyl-[acp] to a propanedioyl~[acp] methyl ester. Each of the one or more cycles of methyl ester shielded carbon chain elongation can include using polypeptides having the activity of one or more (i) a f]-ke(oacy!~[acp] synthase or β- ketothiolase, (it) a 3-oxoacyl-f acp] reductase, ceioacetyl-CoA reductase, a 3-hydroxyacyl- CoA dehydrogenase or a 3~hydroxyb tyryl-CoA dehydrogenase, (lis) an enoyl-CoA hydrolase or a 3~hydroxy cyl~[acp] dehydratase, and (iv) an enoyl-facp] reductase or a trans~2~enoyi- CoA reductase to produce heptanedioyl-[acp] methyl ester from propanedioy!-[acpj methyl ester or produce heptanedioyl-CoA methyl ester from propanedioy!-CoA methyl ester.

According to the present disclosure, the methyl group is not removed from the carboxyl-CoA methyl ester or the carboxyl~[aep] methyl ester following the one or more cycles of carbon chain elongation without at least one further enzymatic step. In some embodiments, a polypeptide having the activity of S-adenosyl-L-methionine (SAM) -dependent methyhransferase can add an initial methyl ester shield to propanedioyi- Co A to form propatiedioyl-CoA methyl ester or propanedioyi- acp] to form propanedioyl- [acp] methyl ester. The polypeptide having the activity of S-aderiosy -L-me!hionine (SAM)- dependent methyhransferase can have at least 50%, at least 60%, at least 70%, or at least 85% sequence identity or homology to the amino acid sequence of SEQ ID NO: 52,

A polypeptide having the activity of an esterase can remove a methyl shield from the methyl-protected C21J ₊ 3 building blocks, wherein n is an integer greater than or equal to one (i.e., C;-Ci9 building blocks), such as monornethyl carboxylate, monomethyl carboxylate semialdehyde, and monomethyl aminocarboxylate. A polypeptide having the activity of an esterase can have at least 50%, at least 60%, at least 70%, or at least 85% sequence identity or homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 ,

The two terminal functional groups can be the same (e.g., amine, formyl, carboxyl, or hydroxyl groups) or can be different (e.g., a terminal amine and a terminal carboxyl group; oi ^¬ a terminal hydroxyl group and a terminal carboxyl group).

A polypeptide having the activity of an aminotransferase or a deacetyiase can enzymaticaliy form an amine group. A polypeptide having the activity of an aminotransferase can have at least 50%, at least 60%, at least 70%, or at least 85% sequence identity or homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 , A polypeptide having the activity of a deacetyiase can have at least 50%, at least 60%, at least 70%, or at least 85% sequence identity or homology to the amino acid sequence of any one of SEQ ID NOs: 42-45.

A polypeptide having the activity of 6-hydroxyhexanoate dehydrogenase, 5- hydroxypentanoate dehydrogenase, 4-hydroxybutyrate dehydratase, or alcohol dehydrogenase can enzymaticaliy form a hydroxyl group.

A polypeptide having an activity selected from thioesterase, esterase, aldehyde dehydrogenase, 7~oxoheptanoate dehydrogenase, 6-oxohexanoate dehydrogenase, CoA- iransferase (e.g. glutaconaie CoA transferase), and reversible CoA-llgase (e.g.. reversible succinate-CoA ligase) can enzymaticaliy form a terminal carboxyl group, A polypeptide having the activity of a thioesterase can have at least 50%, at least 60%, at least 70%. or at least 85% sequence identity or homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195.

A polypeptide having the activity of a carboxylate reductase and a polypeptide having the activity of a phosphopantetheinyl transferase can form a terminal aldehyde group as an intermediate in forming the product, A polypeptide having the activity of a carboxylate reductase can have at least 50%, at least 60%, at least 70%, or at least 85% sequence identity or homology to the amino acid sequence of any one of SEQ ID NOs; 25-39 or SEQ ID NOs: 196-215,

Any of the methods described herein can be performed in a recombinant host by fermentation. The host can be subjected to a cultivation strategy under aerobic, anaerobic, micro-aerobic or mixed oxygen/denitrification cultivation conditions. The host can be cultured under conditions of nutrient limitation The host can be retained using a ceramic hollow fiber membrane to maintain a high cell density during fermentation.

In some embodiments, a cultivation strategy is used to achieve anaerobic, micro- aerobic, or aerobic cultivation conditions.

In some embodiments, the cultivation strategy includes limiting nutrients, such as limiting nitrogen, phosphate, or oxygen.

In any of the methods, the host ^' s tolerance to high concentrations of a difunctional product having an odd number of carbon atoms, such as a €5-0: building block, can be improved through continuous cultivation in a selective environment.

In some embodiments, the host may comprise a deletion in hi H. In some embodiments, the host does not express BioH. in some embodiments, the host may comprise a deletion in met,]. In some embodiments, the host does not express Met J.

The principal carbon source fed to the fermentation can derive from bioiogical or non- bioiogical feedstocks, in some embodiments, the biological feedstock is, includes, or derives from, monosaccharides, disaccharides, iignocellulose, hemicellulose, cellulose, lignin, levulinic acid and formic acid, triglycerides, glycerol, fatty acids, agricultural waste, condensed distillers ^' ' solubles, or municipal waste. In some embodiments, the feedstock is not glucose.

In some embodiments, the non-biological feedstock is or derives from natural gas, syngas, CO ₂ /H ₂ , methanol, ethano!, benzoate, non-volatile residue (NVR) or a caustic wash waste stream, from cyclohexane oxidation processes, or a terephthalic acid isophthalic acid mixture waste stream,

This document also features a recombinant host that Includes at least one exogenous nucleic acid encoding one or more of: (i) an S-adenosyl-L-methionin (SAM)-dependent meinyltransferase, (ii) a β-ketoacyl-facpj synthase or a β-ketothiolase, (iii) a 3-oxoacyl-facp] reductase, acetoacetyl-CoA reductase, a 3-hydroxyacyl-CoA dehydrogenase or a 3- hydroxyhutyryl-CoA dehydrogenase, (iv) an enoyl-CoA hydratase or 3-hydroxyacyl-[ cp] dehydratase, and (v) an enoyl~[acp] reductase or a trans~2~enoyl-CoA reductase, said host producing a carboxyl-[acp] methyl ester or a carboxyl-CoA methyl ester.

A recombinant host producing a carboxyi-jacp] methyl ester or a carboxyl-CoA methyl ester can further comprise at least one exogenous nucleic acid encoding one or more of an esterase, a thioesterase, an aldehyde dehydrogenase, a 7-oxoheptanoate dehydrogenase, a 6-oxohexanoate dehydrogenase, a glutaconate CoA~transferase, a reversible s ccinyl-CoA ligase, an acetylating aldehyde dehydrogenase, or a carboxylate reductase, said host producing a carboxyiic acid, a monomethyl carboxylate, a carboxylate semialdehyde or a monomethyl carboxylate semialdehyde,

A recombinant host producing a carboxylate semialdehyde can further comprise at least one exogenous nucleic acid encoding an aminotransferase, said host producing an aminocarboxyiate.

A recombinant host producing a monomethyl carboxylate semialdehyde further can include at least one exogenous nucleic acid encoding an esterase, said host producing a carboxylate semialdehyde,

A recombinant host producing .monomethyl carboxylate further can include at least one exogenous nucleic acid encoding an esterase, said host producing a dicarboxylic acid.

A recombinant host producing monomethyl carboxylate further can include at least one exogenous nucleic acid encoding a carboxylate reductase, optionally in combination with a phosphopantetheine transferase enhancer, said host producing a monomethyl carboxylate semialdehyde,

A recombinant host producing monomethyl carboxylate semialdehyde further can comprise at least one exogenous nucleic acid encoding an aminotransferase, and/or at least- one exogenous nucleic acid encoding an esterase, said host producing an aminocarboxyiate. A recombinant host producing carboxylate semialdehyde further can comprise at least one exogenous nucleic acid encoding a 4-hydroxybutyrate dehydrogenase, a 5- hydroxypentanoate dehydrogenase, or a 6-hydroxyh xanoate dehydrogenase, said host producing a hydroxycarboxylaie.

A recombinant host producing monomethyl carboxylate semiaidehyde further can include at least one exogenous nucleic acid encoding an esterase, and/or a 4-hydroxybutyrate dehydrogenase, a 5-hydroxypentanoate dehydrogenase or a 6-hydroxyhexanoate dehydrogenase, said host producing a hydroxycarboxylate,

A recombinant host producing carboxylate semiaidehyde, an aminoearboxylate, or a hydroxycarboxylaie acid further can comprise at least one exogenous nucleic acid encoding a carboxylate reductase, an aminotransferase, a deacetylase, an N-aeeiyl transferase, or an alcohol dehydrogenase, said host producing a diamine,

A recombinant host producing a hydroxycarboxylate further can further comprise at least one exogenous nucleic acid encoding a carboxylate reductase or an alcohol dehydrogenase, said host producing a diol.

A recombinant host producing an aminoearboxylate further can include at least one exogenous nucleic acid encoding a carboxylate reductase or an alcohol dehydrogenase, the host producing an aminoaldehyde.

The recombinant host can be a prokaryote, e.g., from the genus Escherichia such as Escherichia coli; from the genus Clostridia such as Clostridium Ijungdahlii, Clostridium autoethanogenum or Clostridium kluyverv, from the genus Corynehacteria such as Corynebacteri m glutamicum; from the genus Cupriavidus such as Cupriavidus necator or Cupriavidus metallidurans; from the genus Pseudomonas such as Pse domonas fluorescens, Pseudomonas putida or Pseudomonas o!eavora ; from the genus Delftia acidovorans; from the genus Bacillus such as Bacillus suhtillis; from the genes Lactobacillus such as Lactobacillus delbrueckii; from the genus Lactococcus such as Lactococcus laciis; or from the genus Rhodococcus such as Rhodococcus equi. In some embodiments, the host is not Escherichia coli.

^' [ ^' he recombinant host can be a eukaryote, e.g.. a eukaryote from the genus Aspergillus such as Aspergillus niger; from the genus Saccharomyces such as Saccharomyces cerevisiae; from the genus Pichia such as Pichia pastoris; from the genus Yarrowia such as Yarrowia lipolytics, from the genus Iss tchenkia such as Jssathenkia orientalis, from the genus Debaryomyces such as Debaryomyces hansenii, from the genus Arxula such as Arxula adenoinivorans, or from the genus Kluyveromyces such as Kluyveromyces lactis.

Any of the recornbinani hosts described herein further can comprise one or more of the following attenuated enzymes: polyhydroxyalkanoate synthase, an acetyl-CoA thioesterase, an acetyl-CoA specific β-ketothioiase, a phosphotransacetylase forming acetate, an acetate kinase, a lactate dehydrogenase, a men quinol-fum rate oxidoreductase, a 2- oxoacid decarboxylase producing isobutanoi, an alcohol dehydrogenase forming ethanol, a triose phosphate isomerase, a pyruvate decarboxylase, a glucose-6-phosphate isomerase, a transhydrogenase dissipating the NADH or NADPH imbalance, an glutamate dehydrogenase dissipating the NADH or NADPH imbalance, a NADH/NADPH-utilizing glutamate dehydrogenase, a pimeloyl-CoA dehydrogenase; an acyl-CoA dehydrogenas accepting C7 building blocks and central precursors as substrates; a glutaryl-CoA dehydrogenase; and/or a pimeloyl-CoA synthetase.

As used herein, "attenuation" refers to downreguiation or inactivation of gene expression.

Any of the recombinant hosts described herein further can overexpress one or more genes encoding: an acetyl-CoA synthetase, a 6-phosphogluconate dehydrogenase; a transketolase; a puridine nucleotide transhydrogenase; a formate dehydrogenase: a glyceraldehydeSP-dehydrogenase; a w /i ^' c enzyme; a glucose-6-phosphate dehydrogenase; a fructose 1,6 dlpkospkatase; a L-alanine dehydrogenase; a L-glutamate dehydrogenase specific to the NADH or NADPH used to generate a co-factor imbalance; a methanol dehydrogenase, a formaldehyde dehydrogenase, a diamine transporter; a dicarboxylate transporter an S-adenosylmethionine synthetase; and/or a multidrug transporter,

Any of the recombinant hosts described herein may comprise a deletion in In some embodiments, the recombinant host does not express BioH. in some embodiments, the recombinant host may comprise a deletion in met J. in some embodiments, the recombinant host does not express Met J.

The reactions of the pathways described herein can be performed in one or more ceil (e.g., host cell) strains (a) naturally expressing one or more relevant enzymes, (b) genetically engineered to express one or more relevant enzymes, or (c) naturally expressing one or more relevant enzymes and genetically engineered to express one or more relevant enzymes. Alternatively, relevant enzymes can be extracted from any of the above types of host cells and used in a purified or semi-purified form. Extracted enzymes can optionally be immobilized to a solid substrate such as the floors and/or walls of appropriate reaction vessels. Moreover, such extracts include lysates (e.g. ceil l sates) that can be used as sources of relevant enzymes, in the methods provided by the document, all the steps can be performed in cells (e.g., host cells), ail the steps can be performed using extracted enzymes, or some of the steps can. be performed in cells and others can be performed using extracted enzymes.

Many of the enzymes described herein catalyze reversible reactions, and the reaction of interest may be the reverse of the described reaction. The schematic pathways shown in FIGs. 1 -8 illustrate reactions of interest for each of the intermediates, wherein n cycles of methyl ester shielded carbon chain elongation occur, wherein n is an integer greater than or equal to one. Example schematics shown in FIGs. 33-40 are specific to C (« ^:::: 2).

In some embodiments, the host microorganism's endogenous biochemical network is attenuated or augmented to (1) ensure the intracellular availability of 2-oxoglutarate and 2- oxoadipate, (2) create an NAD ⁺ imbalance that may only be balanced via the formation of a difunetional product having an odd number of carbon atoms (i.e., a C5- 9 building block), (3) prevent degradation of central metabolites, central precursors leading to and including difunetional products having an odd number of carbon, atoms (i.e., C5-C19 building blocks) and (4) ensure efficient efflux from the cell.

Unless otherwise defined, all technical and scientific terms used herei have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although methods and materials similar or equivalent to those described herein can. be used to practice the invention, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety, in case of conflict, the present specification, including definitions, will control In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.

The details of one or more embodiments of the invention are set forth in the accompanying drawings and the description below. Other features, objects, and advantages of the invention will be apparent from the description and the drawings, and from the claims. The word "comprising" in the claims may be replaced by "consisting essentially of or with "consisting of," according to standard practice in patent law.

Embodiments of the disclosure include:

1. A method for biosynthesizing a difunetioiiai product having an odd number of carbon atoms in vitro or in a recombinant host, said method comprising:

enzymatically synthesizing an aliphatic carhon chain backbone having an odd number of carbon atoms from (i) acetyl-CoA and propanedioyl-CoA via one or more cycles of methyl ester shielded carbon chain elongation or (ii) propanedioyl-facp] via one or more cycles of methyl ester shielded carbon chain elongation;

enzymatically forming a first terminal functional group selected from carboxyl, amine, formyL and hydroxy! groups in said backbone while maintaining said methyl ester shield for at least one further enzymatic step: and

enzyrnatically forming a second terminal functional group selected from carboxyl. amine, formyl, and hydroxyl groups in said backbone, thereby forming said difunetional product.

2. The method of embodiment 1 , wherein each of said one or more cycles of carbon chain elongation comprises using (i) a polypeptide having the activity of a β-ketoacyl-facp] synthase or a β-ketotkiolase, (ii) a polypeptide having the activity of a 3-oxoacyl~[ cp] reductase, an aceto cetyl~CoA reductase, a 3-hydroxy cyl~CoA dehydrogenase, or a 3- hydroxyb tyryl-CoA dehydrogenase, (iii) an enoyl-CoA hyd tase or a 3-hydroxyacyl-[acp] dehydratase, and (iv) an enoyl-[acp] reductase or a trans-2~enoyl~CoA reductase.

3. The method of embodiments 1 or 2, wherein said difunetional product has at least five carbon atoms.

4. The method of embodiment 3, wherein said difunetional product has five, seven, nine, eleven, thirteen, fifteen, seventeen, or nineteen carbon atoms.

5. The method of embodiment 3, wherein said difunetional product has five, seven, nine, seventeen, or nineteen carhon atoms. 6. The method of embodiment 1, wherein said aliphatic carbon chain backbone is i) pentanedioyl-[acp] methyl ester or pentanedioyl-CoA methyl ester, ii) heptanedioyl-[acp] methyl ester or heptanedioyl-CoA methyl ester, iii) nonanedioyl~[aep] methyl ester or nonanedioyl-CoA methyl ester, iv) ndecanedioyl- acp] methyl ester or undecanedioyl-CoA methyl ester, v) tridecanedioyh[acp] methyl ester or trldecanedioyi-CoA methyl ester, vi) pentadeeanedioyl-[aep] methyl ester or pentadeeanedioyl-CoA methyl ester, vii) heptadecanedioyh[acp] methyl ester or heptadecanedioyl-CoA methyl ester, or viii) nonadecanedioyl- [acp j methyl ester or nonadecanedioyl-CoA methyl ester,

7. The method of any one of embodiments 1 to 6, wherein a polypeptide having the activity of a S-adenos l-L-methionine (SAM) -dependent methyliransferase converts propanedioyl-CoA to propanedioyi-CoA methyl ester or converts propa.nedioyl~[acp] to propanedioyl-[acp] methyl ester before said one or more cycles of methyl ester shielded carbon chain elongation,

8. The method of embodiment 7, wherein the polypeptide having the activity of a S~ adenosyl-L-methionine (SAM)-dependent methyliransferase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52,

9. The method of any one of embodiments 1 to 6, wherein said at least one further enzymatic step comprises the enzymatic conversion of said aliphatic carbon chain backbone to a monomeihyl earboxylate. 10. The method of embodiment 9, wherein said at least on further enzymatic step also produces holo-ACP or holo-CoA.

1 1 , The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of pentanedioyl-[acp] methyl ester to monomethyl pentanedioate or pentanedioyl-CoA methyl ester to monomethyl pentaneclioate. 12. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of heptanedioyl-[acp] methyl ester to monomethy! heptanedioate or heptanedioyl-CoA methyl ester to monomethyl heptanedioate. 13. The method of embodiment 9, wherein said ai least one further enzymatic step comprises the enzymatic conversion of nonanedioyl-[acp] methyl ester to monomethyl nonanedioate or nonanedioyl-CoA methyl ester to monomeihyl nonanedioate.

14. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of undecanedioyl-jacp] methyl ester to monomethyl undecanedioate or undeeanedioyl-CoA methyl ester to monomethyl undeeanedioate,

15. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of trideeanedioyl-[aep] methyl ester to mononiethyi tridecanedioate or tridecanedioyl-CoA methyl ester to monomethyl tride anedioate. 16. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymati conversion of pentadecanedioyl-j aep] methyl ester to monomethyl pentadeeanedioate or pentadecanedioyl-CoA methyl ester to monomethyl pentadecanedioate,

17. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of heptadecanedio yl- [acp] methyl ester to monomethyl heptadecanedioate or heptadecanedioyl-CoA methyl ester to monomethyl heptadeeanedioate.

18. The method of embodiment 9, wherein said at least one further enzymatic step comprises the enzymatic conversion of nonadeca.nedioyl-[acp] methyl ester to monomethyl nonadecanedioate or nonadeeanedioyl-Co A methyl ester to monomethyl nonadecanedioate.

19. The method of any of embodiments 11 to 18, wherein a polypeptide having the activity of a thioesterase enzymatically forms said monomethyl pentanedioate. monomethyl heptanedioate, monomethyl nonanedioate. monomethyl undecanedioate, monomethyl tridecanedioate, monomethyl pentadeeanedioate, monomethyl heptadecanedioate, or monomethyl nonadecanedioate; and either holo-ACP or holo-CoA.

20. The method of embodiment 19, wherein said polypeptide having the activity of a thioesterase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID Os: 58 13 or SEQ ID NOs: 182-195. 21. The method of embodiment 9, wherein said at leas one farther enzymatic step further comprises the enzymatic conversion of said monomefhyl carboxylate to a monomethyi carboxylate semialdehyde,

22. The method of embodiment 21, wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi pentanedioate to methyl 5- oxopentanoate.

23. The method of embodiment 21, wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi heptanedioate to methyl 7- oxoheptanoate, 24, The method of embodiment 21 , wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi nonanedioate to methyl 9-oxononanoate.

25, The method of embodiment 21 , wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi undecanedioate to methyl 11 - oxotmdecanoate, 26, The method of embodiment 21, wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi tridecanedioate comprises methyl 13- oxotridecanoate.

27. The method of embodiment 21, wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi pentadecaneclioate comprises methyl 15- oxopentadeeanoate.

28. The method of embodiment 21 , wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi heptadecanedioate to methyl 17- oxoheptadecanoate.

29. The method of embodiment 21 , wherein said at least one further enzymatic step comprises the enzymatic conversion of monomethyi nonadecanedioate to methyl 19- oxononadecanoate. 30. The method of embodiment 21 , wherein a polypeptide having the activity of a carboxylate reductase enzymatieai!y forms said monomethyl carboxylate semialdehyde.

31. The method of embodiment 30, wherein said polypeptide having the activity of a carboxylate reductase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to th amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215.

32. The method of embodiment 21 , wherein said at least one farther enzymatic step further comprises the enzymatic conversion of said monomethyl carboxylate semialdehyde to a monomethyl aminocarboxylate, 33. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 5-oxopentanoate to monomethyl 5- aminopentanoate.

34, The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 7-oxoheptanoate to monomethyl 7- ammoheptanoat .

35. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 9-oxononanoate to monomethyl 9~ aminononan ate,

36. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 1 1 -oxoundecanoate to monomethyl I I- aminoundecanoaie.

37, The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 13-oxotridecanoate to monomethyl 13- aminotridecanoa e. 38. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 15-oxopentadecanoate to monomethyl 15- aminopentadecanoate.

39. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl ] 7-oxohepiadecanoaie to monomethyl 17- aminoheptadecat oate,

40. The method of embodiment 32, wherein said at least one further enzymatic step comprises the enzymatic conversion of methyl 19-oxononadecanoate to monomethyl 19- aminononadecanoaie. 41. The method of embodiment 32. wherein a polypeptide having the activity of an aminotransferase enzymatically forms said monomethyl aminocarboxylate,

42. The method of embodiment 41, wherein, said polypeptide having the activity of an aminotransferase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167481 ,

43. The method of any one of embodiments 1 to 6, wherein said at least one further enzymatic step comprises the enzymatic conversion of said aliphatic carbon chain backbone to a monomethyl carboxylate semialdehyde,

44. The method of embodiment 43, wherein said at least one further enzymatic step also produces holo-ACP or holo-CoA.

45. The method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of penianedioyl-[acp] methyl ester to methyl 5- oxopentanoate or pentanedioyhCoA methyl ester to methyl 5-oxopentanoate,

46. The method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of heptanedioyl-j aep] methyl ester to methyl 7- oxoheptanoate or heptanedioyl-Co A methyl ester to methyl 7-oxoheptanoate. 47. The method of embodiment 43. wherein said at least one further enzymatic step comprises the enzymatic conversion of nonanedioyl-[acp] methyl ester to methyl 9- oxononanoate or nonanedioyl-Co A methyl ester to methyl 9~oxononanoate,

48. The method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of undeeanedioyl-jacp] methyl ester to methyl 11- oxoundecanoate or und canedioyi-CoA methyl ester to methyl 11-oxoundeeanoate,

49. The method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of tridecanedioyl~[acp] methyl ester to methyl !3~ oxotrideeanoate or tridecanedioyl-CoA methyl ester to methyl 13-oxotridecanoate. 50. ^" Hie method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of pentadecanedioyl-[acp] methyl ester to methyl 15- oxopentadecanoate or pentadecanedioyl-CoA methyl ester to methyl 15-oxopentadecanoate.

51. The method of embodiment 43, wherei said at least one further enzymatic step comprises the enzymatic conversion of heptadeeanedioyi-j acp] methyl ester to methyl 17- oxoheptadeeanoate or heptadecanedioyl-CoA methyl ester to methyl ί 7-oxo eptadecanoate.

52. The method of embodiment 43, wherein said at least one further enzymatic step comprises the enzymatic conversion of nonadecanedioyI~[acp] methyl ester to methyl 19- oxononadeeanoate or nonadecanedioy!-CoA methyl ester to methyl 19-oxononadecanoate.

53. The method of embodiment 44. wherein a polypeptid having the activity of an acetylaiirig aldehyde dehydrogenase enzymaticaily forms said monomethy! carboxyiate semialdehyde and either holo-ACF or holo-CoA.

54. The method of embodiment 53, wherein said polypeptide having the activity of an acetylatmg aldehyde dehydrogenase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18 or SEQ ID NO: 19, 55. The method of embodiment 9, wherein a second terminal functional group is formed by the enzymatic conversion of said monomethy! carboxyiate to a dicarboxylic acid, 56, The method of embodiment 55, wherein said dicarboxylic acid is pentanedioic acid, heptanedioic acid, nonaiiedioic acid, undecanedioic acid, trideeanedioie acid, pentadecanedioic acid, heptadeeanedioic acid, or nonadecanedioic acid.

57. The method of embodiment 55, wherein a polypeptide having the activity of an esterase enzymaiiealiy forms said dicarboxylic acid.

5S, The method of embodiment 56, wherein a polypeptide having the activity of an esterase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NO: 50 or SEQ ID NO: 51.

59. The method of embodiment 21 or 43. wherein a second terminal functional group is formed by the enzymatic conversion of said monomethyl carboxyiate semialdehyde to a carboxyiate semia ! dehyd e .

60, The method of embodiment 59, wherein said carboxyiate semialdehyde is 5- oxopentanoate, 7-oxobepianoate, 9-oxononanoate, ί .1 -oxoundecanoaie, 13-oxotridecanoate. 15-oxopentadecanoaie, 17-oxoheptadecanoate, or 19-oxononadeea.noate. 61. The method of embodiment 59, wherein a polypeptide having the activity of an esterase enzymaiiealiy forms said carboxyiate semialdehyde,

62, The method of embodiment 60, wherein said polypeptide having the activity of an esterase has at least 70% sequence identity to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 . 63, The method of embodiment 32, wherein a second terminal group is formed by the enzymatic conversion of said monomethyl aminocarboxylate to an aminocarboxylate.

64, The method of embodiment 63, wherein said aminocarboxylate is 5-ammopentanoate, 7-aminoheptanoate, 9-aminononanoate, l l-aminoundecanoa e, ί 3-aminotridecanoate, 15- aminopentadecanoate, 17~aminoheptadecanoate, or 19~ammononaclecanoaie. 65, The method of embodiment 63, wherein a polypepiide having the activity of an esterase enzymaiiealiy forms said aminocarboxylate. 66. The method of embodiment 65, wherein said polypeptide having the activity of an esterase has at least 50%, at least 60%, at least 70%, or ai least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51.

67. The method of embodiment 59. further comprising enzymatically converting said earboxylate semialdehyde to an aminoearboxylate.

68. The- method of embodiment 67. wherein said aminoearboxylate is 5-amiriopentanoate, 7-aminoheptanoate, 9-aminononanoate, 1 1 -aminoundeeanoate. 13-aminotrideeanoaie, 15- aminopentadeeanoate, 17-arainoheptadecanoate or 19-aminononadecanoate.

69. The method of embodiment 68, wherein an aminotransferase enzymatically forms said aminoearboxylate.

70. The method of embodiment 72, wherein said aminotransferase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs; 167-181.

71. The method of embodiment 59. further comprising enzymatically converting said earboxylate semialdehyde to a hydroxyearboxylate.

72. The method of embodiment 71 , wherein said hydroxyearboxylate Is 5- hydroxypentanoate, 7-hydroxyheptanoate, 9-liydroxynonanoate, 1 1 -hydroxyundecanoate, 13- hydroxytrideeanoate, 15-hydroxypentadecanoate, 17-hydroxyheptadecanoate, or 19- hydroxynonadecanoate. 73, The method of embodiment 71 , wherein a polypeptide having the activity of an alcohol dehydrogenase enzymatically forms said hydroxyearboxylate.

74. The method of embodiment 73, wherein said polypeptide having the activity of an alcohol dehydrogenase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23, 75. The method of embodiment 59, further comprising enzymatically converting said earboxylate semialdehyde to a diamine, 76. The method of embodiment 75, wherein said carboxyiate semi aldehyde is enzyrnatlcaily converted to said diamine in one or more steps involving a polypeptide having the activity of a carboxyiate reductase and a polypeptide having the activity of an aminotransferase, 77. The method of embodiment 76, wherein said polypeptide having the activity of a carboxyiate reductase has at least 50%, at feast 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ I D NOs: 25-39 or SEQ ID NOs: 196-215 and said polypeptide having the activity of an aminotransferase has at least 70% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

78. The method of embodiment 75, wherein said diamine is pentane- ί ,5-diamine, heptane- 1 ,7-diamine, nonane- 1 ,9-diamine, undecane- 1,11 -diamine, trldecane- 1 ,13 -diamine, pentadecane , 15-diamine, heptadecane- 1 , 17-diamine, or nonadecane- 1 , 19-diamine.

79. The method of embodiments 71 , further comprising enzymatic converting said hydroxycarboxylate to a dioL

80. The method of embodiment 79, wherein said hydroxycarboxylate is enzymatically converted to said diol in one or more steps involving a polypeptide having the activity of a carboxyiate reductase and a polypeptide having the activity of an alcohol dehydrogenase,

81. The method of embodiment 79, wherein said diol is 1,5-pentanediol, L7~heptanediol _s L9~nonanediol, 1,11 -undeeanediol, 1 , 13-tridecanediol, 1 , 1 -perstadecanediol, 1,17- heptadecanediol, or 1, 19-nonadecanediol.

82. The method of embodiment 80, wherein said polypeptide having the activity of a carboxyiate reductase has at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-21 Sand said polypeptide having the activity of an alcohol dehydrogenase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23. 83, The method of embodiment 1 , wherein said two terminal functional groups are the same.

84, The method of embodiment 1, wherein said two terminal functional groups are different, 85. The method of embodiment 84, wherein said di functional product comprises a terminal amine and a terminal carboxyi group,

86. The method of embodiment 84, wherein said difunctionai product comprises a terminal formyl group,

87, The method of embodiment 84, wherein said difunctionai product comprises a terminal hydroxy! group and a terminal carboxyi group,

88. The method of embodiment 83, wherein said two terminal functional groups are amine groups.

89, The method of embodiment 83, wherein said two terminal functional groups are hydroxy 1 groups. 90. The method of embodiment 87 or 89, wherein a polypeptide having the activity of a 6- hydroxyhexanoate dehydrogenase, a polypeptide having the activity of a 5-hydroxypentanoaie dehydrogenase, a polypeptide having the activity of a 4-hydwxyhu yrate dehydratase, or a polypeptide having the activity of an alcohol dehydrogenase enzymadcaily forms a hydroxy! group. 91 , The method of embodiment 85 or 87, wherein a polypeptide having the activity of a thioesterase, a polypeptide having the activity of an aldehyde dehydrogenase, a polypeptide having the activity of a. 7-oxoheptanoate dehydrogenase, a polypeptide having the activity of a 6-oxohexanoale dehydrogenase, a polypeptide having the activity of a glutaconaie CoA- transferase, or a polypeptide having the activity of a reversible suecinyl-CoA ligase enzymaticaily forms a terminal carboxyi group. 92. The method of embodiment 91 , wherein said polypeptide having the activity of a thioesterase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195.

93. The meihod of embodiment 85 or 88. wherein a polypeptide having the activity of an aminotransferase or a polypeptide having the activity of a d cetyl se enzymaticaily forms an amine group.

94. The method of embodiment 93, wherein said polypeptide having the activity of an aminotransferase has at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or or SEQ ID NOs: 167-181.

95. The method of any one of embodiments 1 -94, wherein said method is performed in said recombinant host by fermentation.

96. The method of embodiment 95, wherein said recombinant host is subjected to a cultivation strategy under aerobic, anaerobic, micro-aerobic, or mixed oxygen/denitrifi cation cultivation conditions.

97. The method of embodiment 95 or 96, wherein said recombinant host is cultured under conditions of nutrient limitation.

98. The method according to any one of embodiments 95-97, wherein said recombinant host is retained using a ceramic hollow fiber membrane to maintain a high cell density during fermentation.

99. The method of any one of embodiments 95-98, wherein the principal carbon source fed to the fermentation derives from biological or non-biological feedstocks.

100. The method of embodiment 99, wherein the biological feedstock is, or derives from, monosaccharides, disaecharides, iignoce!!u!ose, hemicellulose, cellulose, lignin, !evulinic acid, formic acid, triglycerides, glycerol, fatty acids, agricultural waste, condensed distillers' solubles, or municipal waste. 101. The method of embodiment 99, wherein the non-biological feedstock is, or derives from, natural gas, syngas,€<¾/¾, methanol, ethanol. benzoate, non-volatile residue (NVR) caustic wash waste stream from cyelohexane oxidation processes, or terephthalic acid / isophthaiic acid mixture waste streams. 102, The method of embodiment 99 or 100, wherein the biological feedstock is not, or does not derive from, glucose.

103. The method of any one of embodiments 95-102, wherein the host is a prokaryote _* .

104. The method of embodiment 103, wherein said prokaryote is from a genus selected from Escherichia;, Clostridia, Corynehacteria, Cupriavidus, Pse domonas, Deljiia, Bacillus Lactobacillus, L ctococcus, and Rhodococcus.

105. The method of embodiment 103 or 104, wherein said prokaryote is selected from Escherichia coli, Clostridium Ijungdahli Clostridium auloethanogenum, Clostridium khtyveri, Corynebacterium ghitamicum, Cupriavidus necator, Cupriavidus meiatlidurans, Pseudomonas fluorescens, Pseudomonas putida or Pseudomonas oleavorans, Delfiia acidovorans, Bacillus subtillis, Lactobacillus delbrueckii, Lactococcus lactis, and Rhodococcus equi,

106. The method of any one of embodiments 103-105, wherein said prokaryote is not Escherichia coli.

107. The method of any one of embodiments 95-102, wherein the host Is a eukaryote. 108. The method of embodiment 107, wherein said eukaryote is from a genus selected from Aspergillus such as from the genus Saccharomyces such as; from the genus Pichia. such as Pichia pastor is; from the genus Yarrowia such as, from the genus IssatchenMa such as, from the genus Deb ryomyces such as, from the genus Arxula such as, or from the genus Kluyveromyces such as, 109. The method of embodiment 107 or 108, wherein said eukaryote is selected from Aspergillus niger, Saccharomyces cerevisiae, Yarrowia lipolytics, Issathenki oriental is, Debaryomyces hansenii, Arxula denoinivorans, and Kluyveromyces lactis.

1.10, The method of any one of embodiments 95-109, wherein said recombinant host's tolerance to high concentrations of a difunctional product is improved through continuous cultivation in a selective environment.

1 1 1 . The method of any one of embodiments 95-110, wherein said one or more of the following enzymes is attenuated in said recombinant host: a polyhydroxyalkanoate synthase, an cetyl-CoA thioesterase, an acetyl-CoA specific β-ketolhiolase, a pkosphotransacetylase forming acetate., an acetate kinase, a lactate dehydrogenase, a menaquinoi-fv.marate oxidorediictase, a 2-oxoacid decarboxylase producing isobutanol, an alcohol dehydrogenase forming ethanoL a triose phosphate isomerase, a pyruvate decarboxylase, a glucose~6~ phosphate isomerase, a transhydrogenase dissipating the NADH or NADPH imbalance, an glutamate dehydrogenase dissipating the NADH or NADPH imbalance, a NADH/NADPH- utilizing glutamate dehydrogenase, a pimeloyl-CoA dehydrogenase; an acyl~CoA dehydrogenase accepting C$, C ₇ , C , Cn, C13, C _\ s, Q- ι _. η, or C;o building blocks and central precursors as substrates; a giutaryi-CoA dehydrogenase', or a pimeloyl-CoA synthetase.

1 12. The method of any one of embodiments 95-111, wherein said host overexpresses one or more genes encoding: an acetyl-CoA synthetase, a 6-phosphogluconate dehydrogenase; a transketolase; a pyridine nucleotide iranshydrogenase; a formate dehydrogenase; a glyceraldehyde~3P-dehydrogenase a malic enzyme; a glv.cose-6 -phosphate dehydrogenase; a fructose 1,6 diphosphatase; a L-alanine dehydrogenase; a. L~glutarna(e dehydrogenase specific to the NADH or NADPH used to generate a co-factor imbalance; a methanol dehydrogenase, a formaldehyde dehydrogenase, diamin transporter; a dicarboxylate transporter; an S-adenosyl eihiomne synthetase; or a multidrug transporter.

1 13. A recombinant host comprising at least one exogenous nucleic acid encoding one or more of: (t) a S-adenosyl-L-methionine (SAM) -dependent me thyltransf erase, (ii) a polypeptide having the activity of a fj~ketoacyi~facp] synthase or a β-ketothiolase, (iii) a polypeptide having the activity of a 3~oxoacyl-[acp] reductase, an aceioacetyl-CoA reductase, a 3- hydroxyacyl-CoA dehydrogenase, or a 3~hydr xybutyryl-CoA dehydrogenase, (iv) an enoyl- CoA hydratase or a 3~}rydroxyacyl~[acp] dehydratase, and (v) an enoyl~[ cp] reductase or a tram-2-enoyl-CoA reductase, said host producing a difunctional product having an odd number of carbon atoms.

1 14. The recombinant host of embodiment 113, said host comprising a deletion in met J,

1 5. The recombinant host of embodiment 113 or 114, wherein said host does not express MetJ.

1 16. The recombinant host of embodiment 113, wherein said host comprises a deletion in bioH.

1 17. The recombinant host of embodiment 113 or 116, wherein said host does not express BioH.

1 18. The recombinant host of any one of embodiments 113-117, said host further comprising at least one exogenous nucleic acid encoding one or more of a thioesterase, an aldehyde dehydrogenase, a 7-oxoheptanoate dehydrogenase, a 6-oxohexanoate dehydrogenase, a glutaconate Co A -transferase, a reversible s ccinyl-CoA ligase, an acetylating aldehyde dehydrogenase, or a carboxylate reductase, said host producing a dicarboxvlic acid having an odd number of carbon atoms.

119. The recombinant host of embodiment 1 18, wherein said dicarboxvlic acid having an odd number of carbon atoms is pentanedioic acid, heptanedioic acid, nonanedioic acid, undecanedioic acid, tridecanedioic acid, pentadecanedioic acid, heptadecanedioic acid, o nonadecanedioic acid.

120. The recombinant host of any one of embodiments 113-117, said host further comprising at least one exogenous nucleic acid encoding an aminotransferase, said host producing an aminocarboxylate having an odd number of carbon atoms.

121. The recombinant host of embodiment 120, wherein said aminocarboxylate having an odd number of carbon atoms is S-aminopcntanoate, 7-arninoheptanoate, 9-aminononanoate. 11 -ammoundecanoate, IS-ammotridecanoate, I S-aminopentadeeanoate, 17- am noheptadecanoate, or 19-aminononadecanoate.

122. The recombinant host of any one of embodiments 113-117, said host further comprising at least one exogenous nucleic acid encoding one or more of a 4 -hydroxy buty rate dehydrogenase, a 5-hydroxypentanoate dehydrogenase or a 6-hydroxyhexanoate dehydrogenase, said host producing a hydroxycarboxylate having an odd number of carbon atoms.

123. The recombinant host of embodiment 122, wherein said hydroxycarboxylate having an odd number of carbon atoms is 5-hydroxypentanoate. 7-hydroxyheptanoate, 9- hydroxynonanoate, 11-hydroxyundecanoate, 13-hydroxytridecanoate. 15- hydroxypentadecanoate, 1 T-hydroxyheptadecanoate, or 19-hydroxynonadeeanoate.

124. The recombinant host of any one of embodiments 113-123, said host further comprising at least one exogenous nucleic acid encoding one or more of an aminotransferase, a deacetylase, an N-acetyl transferase, or an alcohol dehydrogenase, said host producing a diamine having an odd number of carbon atoms.

125. The recombinant host of embodiment 124, wherein said diamine having an odd number of carbon atoms is pentane-l,5~diamine, heptane- 1 ,7-d.iamine, nonane-l ,9-diamine, undecane- 1 ,11 -diamine, tridecane- 1 , 13-diamine, pentadecane- 1 , 15-diamine, heptadecane- 1,17-diamine, or nonadeeaiie-i,19-diamme, 126. The recombinant host of embodiment 122, said host further comprising at least one exogenous nucleic acid encoding one or more of a (i) carhoxylate reductase enhanced by a phosphopantetheinyl transferase or (ii) an alcohol dehydrogenase, said host producing a diol having an odd number of carbon atoms.

127. The recombinant host of embodiment 126, wherein said diol having an odd number of carbon atoms is 1 ,5-pentanediol, 1,7-heptanedioi, l,9~nonanediol, 1 ,11-undecanediol, 1 ,13- tridecanediol, 1 ,15-pentadecanediol, l,r7-heptadecanediol, or 1 ,19-nonadeeanediol 12S. A non -naturally occurring organism comprising at bast one exogenous nucleic acid encoding at least one polypeptide having the activity of at least one enzyme depicted in any one of FIGs, 1-8. i 29. A nucleic acid construct or expression vector comprising at least one polynucleotide encoding one or more polypeptides having an enzymatic activity, wherein the at least one polynucleotide is operably linked to one or more heterologous control sequences that direct production of the one or more polypeptides, wherein the one or more polypeptides is selected from: (a) a polypeptide having the activity of a 3 ^' -hydroxy >acyl-[acp] dehydratase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 ; (b) a polypeptide having the activity of a 3-kydroxyacyl-CoA dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 2-4; (c) a polypeptide having the activity of a 3-oxoacyl~[ cp ] reductase having at least 50%, at least 60%, at least 70%, o at least 85%> sequence homology to the annuo acid sequence of SEQ ID NO: 5; (d) a polypeptide having the activity of a -hydroxy 'b t rate dehydrogenase having at least 50%, at least 60%, at. least 70%, or at ieast 85% sequence homology to the amino acid sequence of SEQ ID NO: 23: (e) a polypeptide having the activity of a 5-hydroxypen!anoate

dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ 113 NO: 21 ; (f) a polypeptide having the activity of a 6-hydroxyh xanoate dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8; (g) a

polypeptide having the activity of a 6~oxohex no te dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10; (h) a polypeptide having the activity of a 7-oxokeptanoate dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 11 -13; (i) a polypeptide having the activity of a β-ketoacyi-facp] synthase having at least 50%, at least 60%, at Ieast 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NOs: 14-16; (j) a polypeptide having the activity of a β-ketoihlolase having at ieast 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 17; (k) a polypeptide having the activity of an acetylating aldehyde dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ) ID NO: 18 or SEQ ID NO: 19; (i) a polypeptide having the activity of an alcohol dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23; (m) a polypeptide having the activity of an aldehyde dehydrogenase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24; (n) a polypeptide having the activity of a carhox l te reductase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 1 6-215; (o) a polypeptide having the activity of a CoA~transferase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequenc of SEQ ID NO: 40 or SEQ ID NO: 41 ; (p) a polypeptide having the activity of a deaceiylase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45; (q) a polypeptide having the activity of an enoyl-facp] reductase having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46 or SEQ ID NO: 6; (r) a poiypepiide having the activity of an enoyl-CoA h dratase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 47-49; (s) a polypeptide having the activity of an esterase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 ; (t) a polypeptide having the activity of a S-adenosyl-L-metkionine (SAM) -dependent me thyltransf erase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52; (u) a polypeptide having the activity of a N-acetyltransferase having at least 50%i, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53; (v) a polypeptide having the activity of a phosphopanteiheine transferase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57; (w) a polypeptide having the activity of a thioesterase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; (x) a polypeptide having the activity of a trans-2-enoyl-CoA reductase having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 114 or SEQ ID NO: 1 15: and (y) a polypeptide having the activity of an aminotransferase having at least 50%, at least 60%, at [east 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID Os: 116-138 or SEQ ID NOs; 167-1 81 ,

130. A composition comprising the nucleic acid construct or expression vector of embodiment 129,

131. A bio-derived, bio-based or fermentation-derived product, wherein said product comprises:

i. a composition comprising at least one bio-derived, bio-based, or fermentation- derived compound produced or biosynthesized according to any one of embodiments 1 -1.13, or any combination thereof,

ii. a bio-derived, bio-based, or fermentation-derived polymer comprising the bio- derived, bio-based, or fermentation-derived composition or compound of i., or any eonibin ati on thereof,

iii a bio-derived, bio-based, or fermentation-derived resin comprising the bio-derived, bio-based, or fermentation-derived compound or bio-derived, bio-based, or fermentation- derived composition of i. or any combination thereof or the bio-derived, bio-based, or fermentation-derived polymer of it. or any combination thereof,

iv, a molded substance obtained by molding the bio-derived, bio-based, or fermentation-derived polymer of ii, or the bio-derived, bio-based, or fermentation-derived resin of iii., or any combination thereof

v. a bio-derived, bio-based, or fermentation-derived formulation comprising the bio- derived, bio-based, or fermentation-derived composition of L, bio-derived, bio-based, or fermentation-derived compound of L bio-derived, bio-based, or fermentation-derived polymer of if, bio-derived, bio-based, or fermentation-derived resin of iii., o bio-derived, bio-based, or fermentation-derived, molded substance of iv, or any combination thereof, or vi. a bio-derived, bio-based, or fermentation-derived semi-solid or a non-semi-solid stream, comprising the bio-derived, bio-based, or fermentation-derived composition of f, bio- derived, bio-based, or fermentation-derived compound of L bio-derived, bio-based, or fermentation-derived polymer of ii., bio-derived, bio-based, or fermentation-derived resin of hi., bio-derived, bio-based or fermentation-derived formulation of v., or bio-derived, bio- based, or fermentation-derived molded substance of iv., or any combination thereof.

132, The method of any one of embodiments 1 -1 12, wherein the product is in the form of a salt or derivative thereof,

DESCRIPTION OF THE DRAWINGS FIG. 1 provide a schematic of an example biochemical pathway leading to a earboxyl- [acp] methyl ester having an odd number of carbon atoms using polypeptides having the activity of one or more NADPH-dependent enzymes and propanedioyl-[acp] as central metabolite following n cycles of methyl ester shielded carbon chain elongation.

FIG. 2 is a schematic of an example biochemical pathway leading to a carboxyl-CoA methyl ester having an odd number of carbon atoms using polypeptides having the activity of one or more NADPH-dependent enzymes and acetyi-CoA and propanedioyl-CoA as central metabolites.

FIG. 3 is a schematic of an example biochemical pathway leading to carboxyl-CoA methyl ester having an odd number of carbon atoms using polypeptides having the activity of one or more NADH-dependent enzymes and acetyl-CoA and propanedioyl-CoA as central metabolites.

FIG. 4 is a schematic of example biochemical pathways leading to a dicarboxyiic acid having an odd number of carbon atoms using a carboxyl-[acp] methyl ester, a carboxyl-CoA methyl ester, or a monomethyl carboxylate semialdehyde as central precursors.

PIG, 5 is a schematic of example biochemical pathways leading to an aminocarboxylate having an odd number of carbon atoms using a carboxyl-CoA methyl ester, a monomethyl carboxylate, a monomethyl carboxylate semialdehyde, a monomethyl aminocarboxylate, or a dicarboxyiic acid as central precursors,

FIGs. 6A and 6B are schematics of example biochemical pathways leading to a diamine having an odd number of carbon atoms using an aminocarboxylate, a hydroxycarboxyiate, or a carboxylate semialdehyde as central precursors.

FIG, 7 is a schematic of example biochemical pathways leading to a hydroxycarboxyiate having an odd number of carbon atoms using a dicarboxyiic acid, a carboxy-CoA methyl ester, a monomethyl carboxylate semialdehyd , or a carboxylate semialdehyde as central precursors.

FIG. 8 Is a schematic of an example biochemical pathway leading to a diol having an odd number of carbon atoms using a hydroxycarhox late as a central precursor.

FIG. 9 illustrates the structures of carhoxyl-ACP methyl esters produced by n cycles of methyl esier shielded carbon chain elongation for n 0, 1 , 2, 3, 4. 5, 6, 7, and 8.

FIG. 10 Illustrates the structures of carboxyl-CoA methyl esters produced by n cycles of methyl ester shielded carbon chain elongation for n ^::: 0, 1 , 2, 3, 4, 5. 6, 7, and 8.

FIG, 11 illustrates the structures of monomethyl carboxylates produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3. 4, 5, 6, 7, or 8.

FIG. 12 illustrates the structures of monomethyl carboxylate semia!dehydes produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG, 13 illustrates the structures of dicarboxy!ic acids produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1 , 2, 3. 4, 5, 6, 7, or 8,

FIG. 1.4 illustrates the structures of monomethyl aminocarboxylates produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded, carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG. 15 illustrates the structures of aminocarboxylates produced from aliphatic backbones having an odd number of carbon atoms following « cycles of methyl ester shielded carbon chain elongation, where « is 1 , 2, 3, 4, 5, 6, 7. or 8.

FIG. 16 illustrates the structures of carboxylate semia!dehydes produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3. 4, 5, 6, 7, or 8.

FIG, 17 illustrates the structures of aminoaldehydes produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8. FIG, 18 illustrates the structures of diamines produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG, 19 illustrates the structures of hydroxycarboxy!ates produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where w is 1, 2, 3, 4, 5, 6, 7, or 8,

FIG. 20 illustrates the structures of hydroxyamines produced from aliphatic backbones having an odd number of carbon atoms follovsdng n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG. 21 illustrates the structures of aeeta idoearboxy fates produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2. 3, 4, 5, 6. 7, or 8.

FIG, 22 illustrates the structures of acetamidoaldehydes produced from aliphatic- backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n ^' 1 , 2, 3, 4, 5, 6, 7, or 8.

FIG, 23 illustrates the structures of acetamkloamines produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8,

FIG. 24 illustrates the structures of dials produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1 , 2, 3, 4, 5, 6, 7, or 8.

FIG. 25 illustrates the structures of hydroxy aldehydes produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG. 26 illustrates the structures of diois produced from aliphatic backbones having an odd number of carbon atoms following n cycles of methyl ester shielded carbon chain elongation, where n is 1 , 2, 3, 4, 5, 6, 7, or 8.

FIG. 27 illustrates the structures of 3~oxG-earhoxyl~ACP methyl esters produced during the m ^th cycle of methyl shielded carbon chain elongation, where m is 1 , 2, 3, 4, 5, 6, 7, or 8. FIG. 28 illustrates the structures of 3-oxo-carboxyl-CoA methyl esters produced during the m ¹ cycle of methyl shielded carbon chain elongation, where m is 1, 2, 3, 4, 5, 6, 7, or 8.

FIG, 29 illustrates the structures of 3-hydroxy-carboxyl-ACP methyl esters produced during the m ^h cycle of methyl shielded carbon chain elongation, where m is 1, 2, 3, , 5, 6, 7, or 8.

FIG. 30 illustrates d e structures of 3-hydroxy-carboxyl-CoA methyl esters produced during the ^lh cycle of methyl shielded carbon chain elongation, where m is 1, 2, 3, 4, 5. 6, 7, or 8,

FIG. 31 illustrates the structures of 2,3~dehydrocarboxyl~ACP methyl esters produced during the m ¹ⁿ cycle of methyl shielded carbon chain elongation, where m is L 2, 3, 4, 5, 6, 7, or g.

FIG. 32 illustrates the structures of 2,3-dehydrocarboxyi-CoA methyl esters produced during the m ^lh cycle of methyl shielded carbon chain elongation, where m is 1, 2, 3, 4, 5, 6, 7, or 8,

FIG. 33 is a schematic of an example biochemical pathway leading to heptanedioyl- [acp] methyl ester using polypeptides having the activity of one or more NADPH-dependent enzymes and propanedioyl-[aep] as central metabolites.

FIG, 34 is a schematic of an example biochemical pathway leading to hep anedioyi- CoA methyl, ester using polypeptides having the activity of one or more NADPH-dependent enzymes and aeetyi-CoA and propanedioyl-CoA as central metabolites,

FIG, 35 is a schematic of an example biochemical pathway leading to heptanedioyl- CoA methyl ester using polypeptides having the activity of one or more NADH-dependent enzymes and acetyl-CoA and propanedioyl-CoA as central metabolites.

FIG, 36 is a schematic of example biochemical pathways leading to heptanedioate using heptanedioyl-[acp] methyl ester, heptanedioyi -CoA methyl ester, or methyl 7- oxoheptanoate as central precursors.

FIG, 37 is a schematic of example biochemical pathways leading to 7- aminoheptanoate using heptanedioyl-CoA methyl ester, monomethyl heptanedioate, methyl 7- oxoheptanoate, monomethyl 7-aminoheptanoate, or heptanedioate as central precursors. FIG. 38 is a schematic of exemplary biochemical pathways leading to heptane- 1 ,7- diamine using 7-aniinohepianoate, 7~hydroxyheptanoate or 7-oxoheptanoate as central precursors.

FIG. 39 is a schematic of exemplary biochemical pathways leading to 7- hydroxyheptanoate using heptanedioate. heptanedioyl-CoA methyl ester, methyl 7- oxoheptanoate, or 7-oxoheptanoate as central precursors.

FIG. 40 is a schematic of an exemplary biochemical pathway leading to 1 ,7- heptanediol using 7-hydroxyheptanoate as a central precursor.

FIG. 41 is a bar graph summarizing the change in absorhance at 412 rim, which is a measure, of the release of holo-ACP and the activity of thioesterases for converting heptanedioyl~[acp] or heptanedioyl~[acp] methyl ester.

FIG. 42 is a bar graph summarizing the change in absorhance at 340 m after 20 minutes, which is a measure of the consumption of NADPH and activity of carhoxyloie reductases relative to the enzyme only controls (no substrate).

FIG. 43 is a bar graph of the change in absorhance at 340 nm after 20 minutes, which is a measure of the consumption of NADPH and the activity of carhoxylate reductases for converting heptanedioate to 7-oxoheptanoate relative to the empty vector control.

FIX}. 44 is a bar graph of the change in absorhance at 340 nm after 20 minutes, which is a measure of the consumption of NADPH and the activity of carhoxylate reductases for converting 7-hydrox.yheptai5.oate to 7-hydroxyheptanal relative to the empty vector control,

FIG. 45 is a bar graph of the change in absorhance at 340 nm after 20 minutes, which is a measure of the consumption of NADPH and the activity of carhoxylate reductases for converting N7-acetyl-7-aminoheptanoate to N7-acetyl-7-aminoheptanal relative to the empty vector control.

FIG. 46 is a bar graph of the change in absorhance at 340 nm after 20 minutes, which is a measure of the consumption of NADPH and activity of carboxyiate reductases for converting 7-oxoheptanoate to heptanedial relative to the empty vector control,

FIG. 47 is a bar graph summarizing the percent conversion after 4 hours of pyruvate to L-alanine (mol/mol) as a measure of the aminotransferase activity of the enzyme only controls (no substrate). FIG. 48 is a bar graph of the percent conversion after 4 hours of pyruvate to L-alanine (mol/mol) as a measure of the aminotransferase activity for converting 7-aminoheptanoate to 7-oxoheptanoate relative to the empty vector control.

FIG, 49 is a bar graph of the percent conversion after 4 hours of L-alanine to pyruvate (mol/mol) as a measure of the aminotransferase activity for converting 7-oxoheptanoate to 7- aminoheptanoate relative to the empty vector control.

FIG, 50 is a bar graph of the percent conversion after 4 hours of pyruvate to L-alanine (mol/mol) as a measure of the aminotransferase activity for converting heptane- 1,7-diamine to 7-aminoheptanal relative to the empty vector control.

FIG. 51 is a bar graph of the percent conversion after 4 hours of pyruvate to L-alanine

(mol/moi) as a measure of the aminotransferase activity for converting N7-acetyl-l ,7- diaminoheptane to N7-acetyl~7~ammoheptanal relative to the empty vector control,

FIG. 52 is a bar graph of the percent conversion after 4 hours of pyruvate to .-alanine (mol/moi) as a measure of the aminotransferase activity for converting 7-aininoheptanol to 7- oxoheptanol relative to the empty vector control.

FIG. 53 illustrates representative factors and pathways involved in the S-Adenosyl- methionine (SAM) cycle.

FIG. 54 illustrates the genetically modified Escherichia coli (K12) strains used in Example 11.

FIG. 55 illustrates the assay conditions and comments for using genetically modified

Escherichia coli (K12) strains for heptanedioic acid production in shake flask experiments,

FIG. 56 illustrates the assay conditions and comments for using genetically modified Escherichia coli (K12) strains for heptanedioic acid production in shake flask experiments. Compared to FIG. 55, methionine was added in this assay repeat,

FIG. 57 is a bar graph of the production of heptanedioic acid in genetically modified

Escherichia coii (K12) strains having one or more modifications in proteins involved in the initial step of adding a methyl shield to propanedioyl-CoA or propanedioyl~[acp].

FIG. 58 illustrates the HPLC method used to determine the SAM/SAH levels.

FIG. 59 is a bar graph of the production of SAM/g wet ceil weight in genetically modified Escherichia coli (K12) strains having one or more modifications in proteins involved in the initial step of adding a methyl shield to propanedioyi~CoA or propanedioyl- [aep ^' j.

FIG. 60 illustrates plasmids used for genetically modified Escherichia coli strains for in vivo thioesterase activity screens, DETAILED DESCRIPTION

This document provides enzymes, non-natural pathways, cultivation strategies, feedstocks, host microorganisms, and attenuations to the host's biochemical network, which generate an aliphatic carbon chain backbone having an odd number of carbon atoms from central metabolites in which two iemiinai functional groups may be formed leading to the synthesis of difunctional products having an odd number of carbon atoms, such as, for example, dicarboxylic acids, carboxylate semiaidehydes, aminoearboxylates, hydroxycarboxylates, diamines, and diols. In some embodiments, the difvmciional products have tlve, seven, nine, eleven, thirteen, fifteen, seventeen, or nineteen carbon atoms (i.e., Cs~ Cj building blocks),

As used herein, a "bio-based product" is a product in which both the feedstock (e.g., sugars from sugar cane, corn, wood; biomass; waste streams from agricultural processes) and the conversion process to the product are biologically based (e.g., fernientation/enzyrnatic transformation involving a biological host/organism/enzyme). As used herein, a ""bio-derived product" is a product in which one of the feedstocks (e.g. , sugars from sugar cane, corn, wood; biomass; waste streams from agricultural processes) or the conversion process to the product is biologically based (e.g. , fermentation/enzymatic transformation involving a biological host/organism/enzyme).

As used herein, a "fermentation-derived product" is a product produced by fermentation involving a biological host or organism.

As used herein, the term building block" denotes a carbon chain aliphatic backbone having (2n÷3) carbon atoms and two terminal functional groups, wherein n is an integer greater than or equal to one, such as a carboxyl-[acp] methyl ester or a carboxyl-CoA methyl ester. For example, as used herein, the term "C _s building block" denotes a difunctional product having a five (5) carbon chain aliphatic backbone. For example, as used herein, the term "C _? building block" denotes a difunctional product having a seven (7) carbon chain aliphatic backbone. For example, as used herein, the term "C¾ building block" denotes a difunctional product having a nine (9) carbon chain aliphatic backbone. For example, as used herein, the term "Cn building block" denotes a difunctional product having an eleven (1 1) carbon chain aliphatic backbone. For example, as used herein, the term "C _{3 building block." denotes a difunctional product having a thirteen (13) carbon chain aliphatic backbone. For example, as used herein, the term "Cu building block" denotes a difunctional product haying a fifteen (15) carbon chain aliphatic backbone. For example, as used herein, the term "On building block" denotes a difunctional product having a seventeen (17) carbon chain aliphatic backbone. For example, as used herein, the term "Q building block'" denotes a difunctional product having a nineteen (19) carbon chain aliphatic backbone,

As used herein, the term "C5-C19 building block" means a building block selected from a€5 building block, a C? building block, a i¾ building block, a Cu building block, a Cm building block, a C15 building block, a Cn building block, or a C 9 building block.

As used herein, the term "central precursor" is used to denote any metabolite in any metabolic pathway shown herein leading to the synthesis of a difunctional product having an odd number of carbon atoms, e.g., a Cj-C; _? building block. In some embodiments, a C5-C19 building block may serve as a central precursor for the synthesis of another C5-C19 building block.

The term "central metabolite" is used herein to denote a metabolite that is produced in all microorganisms to support, growth.

Host microorganisms described herein can include endogenous pathways that can be manipulated, such that one or more difunctional products having an. odd number of carbon atoms (e.g., a C5-C19 building block) can be produced. In an endogenous pathway, the host microorganism naturally expresses all of the enzymes catalyzing the reactions within the pathway, A host microorganism containing an engineered pathway does not naturally express all of the enzymes catalyzing the reactions within the pathway but has been engineered such that all of the enzymes within the pathway are expressed, in the host.

The term "exogenous" as used herein with reference to a nucleic acid (or a protein) and a host refers to a nucleic acid that does not occur in (and cannot be obtained from) a cell of that particular type as it is found in nature or a protein encoded by such a nucleic acid. Thus, a non-naturally-occ rnng nucleic acid is considered to be exogenous to a host once In the host. It is important to note that non-naturally-occurring nucleic acids can contain nucleic acid subsequences or fragments of nucleic acid sequences that are found in nature provided the nucleic acid as a whole does not exist in nature. For example, a nucleic acid molecule containing a genomic DNA sequence within an expression vector is non~naiurally-oeeurring nucleic acid, and thus is exogenous to a host cell once introduced into the host, since that nucleic acid molecule as a whole (genomic DNA plus vector DNA) does not exist in nature, Thus, any vector, autonomously replicating plasmid, or vims (e.g., retrovirus, adenovirus, or herpes virus) that as a whole does not exist in nature is considered to be non-naturally- occurring nucleic acid. It follows that genomic DNA fragments produced by PGR or restriction endonuclease treatment as well as cDNAs are considered to he non-naturally- occurring nucleic acids since they exist as separate molecules not found in nature, It also follows that any nucleic acid containing a promoter sequence and polypeptide-encoding sequence (e.g., cDNA or genomic DNA) in an arrangement not found in nature is a non- naturally-occurring nucleic acid. A nucleic acid that is naturally-occurring can be exogenous to a particular host microorganism. For example, an entire chromosome isolated from a cell of yeast x is an exogenous nucleic acid with respect to a cell of yeast y once that chromosome is introduced into a cell of yeast y,

In contrast, the term "endogenous" as used herein with reference to a nucleic acid (e.g., a gene) (or a protein) and a host refers to a nucleic acid (or protein) that does occur in (and can he obtained from) that particular host as it is found in nature. Moreover, a cell "endogenouslv expressing" a nucleic acid (or protein) expresses that nucleic acid (or protein) as does a host of the same particular type as it is found in nature. Moreover, a host "endogenously producing" or that "eiidogenously produces" a nucleic acid, protein, or other compound produces that nucleic acid, protein, or compound as does a host of the same particular type as it is found in nature.

For example, depending on the host and the compounds produced by the host, one or more polypeptides having the activity of one or more the following enzymes may be expressed in the host in addition to an S-adenosyl-L-methionine (SAM) -dependent methyltransferase : a β-keto cyl-f acp] synthase, a β-ketothiolase, a 3~oxoacyl-[acp] reductase, aeetoaceiyl-CoA reductase, a 3-hydroxy cyl-CoA dehydrogenase, a 3-hydroxybuiyryl-CoA dehydrogenase, an enoyl-CoA hydratase, 3-hydroxyacyl-[acp] dehydratase, an enoyl-f cp] reductase, a trans-2-enoyi-CoA reductase, an esterase, a thioesterase, a reversible Gt¾4 Ugase, a CoA-transferase, an acetylaiing aldehyde dehydrogenase, a 6-oxohexanoate dehydrogenase, a 7-oxoheptanoate dehydrogenase, an aldehyde dehydrogenase, a carhoxylate reductase, an aminotransferase, a N~acetyl transferase, an alcohol dehydrogenase, a deacetyiase, a 6- hydroxyhexanoaie dehydrogenase, a 5-hydroxypentanoate dehydrogenase, or a - hydroxyh tyraie dehydrogenase. In recombinant hosts expressing a polypeptide having the activity of a carhoxylate reductase, a polypeptide having the activity of phosphopantetheinyl transferase also can be expressed to enhance the activity of the polypeptide having the activity of a carhoxylate reductase.

For example, a recombinant host can include at least one exogenous nucleic acid encoding one or more of (i) an S-adenosyl-L~methionine (SAM)-dependen! methyltransferase, (ii) a β-ketoacyl-facp] synthase or a β-ketothiolase, (iii) a 3-oxoacyl~[acp] reductase, acetoacetyl-CoA reductase, a 3-hydroxyacyl-CoA dehydrogenas or a 3-hydroxybutyryl~CoA dehydrogenase, (ivj an enoyl-CoA hydratase or 3-hydroxyacyl-[acp] dehydratase, (v) an enoyl- acpj reductase or a trans-2-enoyl-CoA reductase nd produce a carboxyl-[acp] methyl ester or a carboxyl-CoA methyl ester. In some embodiments, the carboxyl-jaep] methyl ester or carboxyl-CoA methyl ester is: i) pentanedioyl-facp] methyl ester or pentanedioyl-CoA methyl ester, ii) heptanedioyi~[acp] methyl ester or heptanedioyl-CoA methyl ester, iii) nonanedioyl-[aep] methyl ester or nonanedioyl-CoA methyl ester, iv) undecanedioyl-jaep] methyl ester or undecanedioyl-CoA methyl ester, v) tridecanedioyl-[acp] methyl ester or tridecanedioyl-CoA methyl ester, vi) pentadecanedioyl~[acp methyl ester or pentadecanedioyl-CoA methyl ester, vii) heptadecanedioyl~[acp] methyl ester or heptadecanedioyi-CoA methyl ester, or viii) nonadeeanedioyl~[acp] methyl ester or nonadeeanedioyl CoA methyl ester.

Such recombinant hosts producing a carhoxyl-[aep] methyl ester or a carboxyl-CoA methyl ester farther can include at least one exogenous nucleic acid encoding one or more of an esterase, a thioesterase, an aldehyde dehydrogenase, a 7-oxoheptanoate dehydrogenase, a 6-oxohexanoate dehydrogenase, a ghitaconate CoA-transferase, a reversible succinyl-CoA ligase, an acetylaiing aldehyde dehydrogenase, or a carhoxylate reductase and produce a diearboxylic acid, monomethy! carhoxylate. carhoxylate sernialdehyde or rnonomethyl carhoxylate sernialdehyde. For example, a recombinant host producing a carboxyl-jaep] methyl ester or a carboxyl-CoA methyl ester further can include at least, one exogenous nucleic acid encoding one or more of a thioesterase, a reversible CoA-ligase (e.g., a reversible succinyl-CoA ligase), or a CoA transferase (e.g., a glutaconaie CoA-transferase) and produce a monomethyl carboxvlate, For example, a recombinant host producing a monomethyl carboxvlate further can include at least one exogenous nucleic acid encoding an esterase and produce a dicarboxylic acid. For example, a recombinant host producing a carboxyl-CoA methyl ester further can include at least one exogenous nucleic acid encoding an acetylating aldehyde dehydrogenase and produce a monomethyl carboxvlate semialdehyde. For example, a recombinant host producing a monomethyl carboxvlate semialdehyde further can include at least one exogenous nucleic acid encoding a 7-oxohepkinoaie dehydrogenase or an aldehyde dehydrogenase and produce a monomethyl carboxvlate. For example, a recombinant host producing a monomethyl carboxylaie further can include at least one exogenous nucleic acid encoding an esterase and produce a dicarboxylic acid.

A recombinant host producing a monomethyl carboxylaie semialdehyde further can include ai least one exogenous nucleic acid encoding an aminotransferase and produce a monomethyl aminocarboxylate, A recombinant host producing a monomethyl aminocarboxylate further can include at least one exogenous nucleic acid encoding an esterase and produce an aminocarboxylate. In some embodiments, a recombinant host producing a carboxyl-CoA methyl ester includes at least one exogenous nucleic acid encoding one or more of an esterase, a carboxylaie reductase, and an aminotransferase to produce an aminocarboxylate.

A recombinant host producing a dicarboxylic acid or a carboxylaie semialdehyde further can include at least one exogenous nucleic acid encoding one or more of a 6~ hydroxyhexanoaie dehydrogenase, a 5~hydroxypentanoaie dehydrogenase, or a 4~ hydraxyh ulyr ale dehydrogenase, and produce a hydroxycarboxylate. In some embodiments, a recombinant host producing a carboxyl-CoA methyl ester includes at least one exogenous nucleic acid encoding one or more of an esterase, an acetylating aldehyde dehydrogenase, a 6-hydroxyhexanoate dehydrogenase, a 5-hydroxypenianoale dehydrogenase, or a 4- hydroxyhiilyraie dehydrogenase to produce a hydroxycarboxylate. In some embodiments, a recombinant host producing a dicarboxylic acid includes at least one exogenous nucleic acid encoding one or more of a carboxylaie reductase, a 6-hydroxyhexanoate dehydrogenase, a 5- hydroxypentanoate dehydrogenase, or a 4-hydroxybutyrate dehydrogenase to produce a hydroxycarboxylate

A recombinant host producing an aminocarboxylate, a hydroxycarboxylate, or a carboxylate semiaidehyde further can include at least one exogenous nucleic acid encoding one or more of an aminotransferase, a deacetylase, a N-acetyl transferase, or an alcohol dehydrogenase, and produce a diamine. For example, a recombinant host producing a hydroxycarboxylate can include at least one exogenous nucleic acid encoding one or more of a carboxylate reductase with a phosphopanteiheine transferase enhancer, an aminotransferase, or an alcohol dehydrogenase.

A recombinant host producing a hydroxycarboxylate further can include at least one exogenous nucleic acid encoding a carboxylate reductase with a phosphopanteiheine transferase enhancer and/or an alcohol dehydrogenase, and produce a diol.

A recombinant host producing an am.inocarboxylate further can include at least one exogenous nucleic acid encoding a carboxylate reductase with a phosphopantetheine transferase enhancer and/or at least one exogenous nucleic acid encoding an alcohol dehydrogenase, and produce a hydroxya ine.

Any of the recombinant hosts described herein ma comprise a deletion in biolf a pimelyl-facp] methyl ester esterase classified, for example, under EC 3, 1.1.85. In some embodiments, the recombinant host does not express BioH, In some embodiments, the recombinant host may comprise a step of down-regulating a repressor that inhibits the initial step of adding a methyl ester shield to propanedioyl-CoA or propanedioyl-jacp], S-Adenosyl- methionine (SAM) -dependent meihyitransferases (MTases) catalyze the transfer of methyl groups from SAM to propanedioyl-CoA or propanedioyl-jacp], The met J gene encodes a regulatory protein which when combined SAM represses the expression of the methionine regulon and of enzymes involved in SAM synthesis. Accordingly, in some embodiments, the recombinant host may comprise a deletion in met J, a SAM co-repressor that represses the intial step of adding a methyl ester shield to propanedioyl-CoA or propanedioyl-jacp]. In some embodiments, the recombinant host does not express MetJ. Enzymes

Within an engineered pathway, the enzymes can be from a single source, i.e., from one genus or species, or can be from multiple sources, i.e., different species. Nucleic acids encoding the enzymes described herein have been identified from various organisms and are readily available in publicly available databases such as GenBank, UniProt, or EMBL. Enzyme Commission (EC) numbers for many enzymes are also provided. EC numbers are well known in the art and provide a numerical classification scheme for enzymes based on the chemical reactions they catalyze. An enzyme classified with an EC number to the fourth level is discretely and specifically classified on the basis of the reactions that its members are able to perform, Well known nomenclature databases such as ENZYME, maintained by the Swiss institute of Bioinformatics, and BRENDA provide examples of specific enzymes corresponding to specific EC numbers.

Any of the enzymes described herein that can be used for production of one or more difunctional products having an odd number of carbon atoms (i.e., C5-C19 building blocks) can have at least 50%. at least 60% or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%», at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or at least 100%) to the amino acid sequence of the corresponding wild-type enzyme. It will be appreciated that the sequence identity can be determined on ihe basis of the mature enzyme (e.g., with any signal sequence removed).

The percent identity and homology between two amino acid sequences can be determined as follows. First, the amino acid sequences are aligned using the BLAST 2 Sequences (B12seq) program from the stand-alone version of BLASTZ containing BLASTP version 2.0.14. This stand-alone version of BLASTZ can be obtained from www.fr.com blast or the U.S. government's National Center for Biotechnology Information web site (www.ncbi.nlm.nih.gov), instructions explaining how to use the B12seq program can be found in the readme fde accompanying BLASTZ. BJ2seq performs a comparison between two amino acid sequences using the BLASTP algorithm. To compare two amino acid sequences, the options of B12seq are set as follows: -i is set to a file containing the first amino acid sequence to be compared (e.g., C:\seql.txt); -j is set to a file containing the second amino acid sequence to be compared (e.g., C:\seq2.txt); -p is set to biastp; -o is set to any desired file name (e.g., C:\outpui.ixt); and all other options are left at their default setting. For example, the following command can be used to generate an output file containing a comparison between two amino acid sequences: C:\B12seq -i c:\seql.txt -j c:\seq2.txt -p blastp -o c:\output.txt if the two compared sequences share homology (identity), then the designated output file will present those regions of homology as aligned sequences, if the two compared sequences do not share homology (identity), then the designated output file will not present aligned sequences. Similar procedures can be followed for nucleic acid sequences except that blastn is used.

Second, once aligned, the number of matches is determined by counting the number of positions where an identical amino acid residue is presented in both sequences. The percent identiiy is determined by dividing the number of matches by the length of the full-length polypeptide amino acid sequence followed by multiplying the resulting value by 100. It is noted that the percent identity value is rounded to the nearest tenth. For example, 78.11, 78.12, 78.13, and 78.14 are rounded down to 78.1, while 78.15, 78,16, 78.17, 78.18, and 78.19 are rounded up to 78.2. it also is noted that the length value will always be an integer.

When percentage of sequence identity is usedwith reference to proteins, it is recognized that residue positions which are not identical often differ by conservative amino acid substitutions, where amino acid residues are substituted for other amino acid residues with similar chemical properties (e.g., charge or hydrophobicity) and therefore do not change the functional properties of the molecule. Where sequences differ in conservative substitutions, the percent sequence identity may be adjusted upwards to correct for the conservative nature of the substitution and this process results in "sequence homology" of, e.g., at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%. Means for making this adjustment are well known to those of ordinary skill in the art. Typically, this adjustment involves scoring a conservative substitution as a partial rather than a full mismatch, thereby increasing the percentage sequence identity. Thus, for example, where an identical amino acid is given a score of 1 and a non-conservative substitution is given a score of zero, a conservative substitution is given a score between zero and 1 , The scoring of conservative substitutions is calculated, e.g., according to the algorithm of Meyers and Miller, Computer, Applic. Biol. Sci., 1988, 4, 11-17, e.g., as implemented in the program PC/GENE (Intelligenetics, Mountain View, Cailf., USA), This alignment and the percent homology or identity can be determined using any suitable software program known in the art, for example those described in CURRENT PROTOCOLS IN MOLECULAR BIOLOGY (F. M, Ausubel et al. (eds) 1987, Supplement 30, section 7,7, 18), Such programs may include the GCG Pileup program, FASTA (Pearson et al , Proc. Natl . Acad. Sci. USA, 1988, 85, 2444- 2448), and BLAST (BLAST Manual Aitschui et al, Nat'l Cent, Biotechnol. Inf., Nat'l Lib, Med. (NOB NLM IH), Bethesda, Md„ and Aitschui et al , NAR, 1997, 25, 3389-3402). Another alignment program is ALIGN Plus (Scientific and Educational Software, Pa.), using default parameters. Another sequence software program that finds use is the TFASTA Data Searching Program available in the Sequence Software Package Version 6.0 (Genetics Computer Group, University of Wisconsin, Madison, Wis.),

A conservative substitution is a substitution of one amino acid for another with similar characteristics. Conservative substitutions include substitutions within the following groups; valine, alanine, and glycine: leucine, valine, and isoleucine; aspartic acid and glutamic acid; asparagine and glutamine; serine, cysteine, and threonine; lysine and arginine; and phenylalanine and tyrosine. The nonpolar hydrophobic amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan, and methionine. The polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine, and glutamine, The positively charged (basic) amino acids include arginine, lysine, and histidine. The negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Any substitution of one member of the above mentioned polar, basic, or acidic groups by another member of the same group can be deemed a conservative substitution, By contrast, a non- conservative substitution is a substitution of one amino acid for another with dissimilar characteristics.

For example, a polypeptide having the activity of a 3~hydroxyacyl~[acp] dehydratase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at leas 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 10(3%) to the amino acid sequence of SEQ ID NO; 1.

For example, a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at. least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 2-4.

For example, a polypeptide having the activity of a 3-oxoacyI-[acp] reductase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%. at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 5.

For example, a polypeptide having the activity of a 4-hydroxybutyrate dehydrogenase described, herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 23,

For example, a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 21.

For example, a polypeptide having the activity of a 6-kydroxyhex noate dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least. 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at leas 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 8.

For example, a polypeptide having the activity of a 6-oxohexanoate dehydrogenase described herein can have at least 50%, at least 60%, or at least. 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at. least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10.

For example, a polypeptide having the activity of a 7-oxoheptanoate dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least. 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 11-13.

For example, a polypeptide having the activity of a. β-ketoacy!-f cp] synthase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%>, at least 60%, at least 65%, at leas 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino add sequence of any one of SEQ ID NOs: 14-16.

For example, a polypeptide having the activity of a β-ketothioiase descri bed herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 17.

For example, a polypeptide having the activity of an acety!ating aldehyde dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 18 or SEQ ID NO: 19.

For example, a polypeptide having the activity of an alcohol dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%), at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ D NOs: 20-23.

For example, a polypeptide having the activity of an aldehyde dehydrogenase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 24,

For example, a polypeptide having the activity of a carhoxylate reductase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at leas 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196- 215.

For example, a polypeptide having the activity of a CoA-transferase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ I ^' D NO: 40 or SEQ ID NO: 41.

For example, a polypeptide having the activity of a deacetylase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology {e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at feast 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 42-45,

For example, a polypeptide having the activity of an enoyl-facp] reductase described herein, can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97% _* , at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 46 or SEQ ID NO; 6.

For example, a polypeptide having the activity of an enoyl-CoA hydrolase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least. 85%, at least 90%, at least 95%, at least. 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 47-49.

For example, a polypeptide having the activity of an esterase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51.

For example, a polypeptide having the activity of a S-adenosyl-L-methionine (SAM)- dependent: methyltr nsferase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least

97%, at least 98%, at least 99%, or 100%) to the amino acid sequence of SEQ ID NO: 52.

For example, a polypeptide having the activity of a N-aceiyltransferase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%), at least 70%, at least 75%, at least

80%, at least 85%, at least 90%), at least 95%, at least 97%, at least 98%, at least 99%, or

100%) to the amino acid sequence of SEQ ID NO: 53.

For example, a polypeptide having the activity of a phosphopaniethein transferase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least. 55%, at least 60%, at least 65%, at least 70%, at least

75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98%, at least

99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 54-57.

For example, a polypeptide having the activity of a ihioesterase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least

85%), at least 90%, at least 95%, at least 97%, at least 98%), at least 99%, or 100%) to the amino acid sequence of any one of SEQ ID NOs: 58413 or SEQ ID NOs: 182-195.

Alternative names for a ihioesterase include, but are not limited to, acyl-ACP fnioesterase, acyl-CoA ihioesterase, arylestera.se, lysophospholipase, acyl-[acyl-carrier~protein ] hydrolase, acyl~ACP-kydrolase, acyl-acyl carrier protein hydrolase, oieoyl-ACP ihioesterase, oleoyl-acyl carrier protein ihioesterase, la ryi-acyl-carrier-protein hydrolase, dodecanoyl- acyl-carrier-protein hydrolase, and doaecyl-acyl-carrier protein hydrolase.

For example, a polypeptide having the activity of a trans-2-enoyl-CoA reductase described herein can have at least 50%, at least 60%, or at least 70% sequence identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least

75%, at least 80%, at least 85%., at least 90%, at least 95%, at least 97%, at least 98%, at least

99%, or 100%) to the amino acid sequence of SEQ ID NO: 7, SEQ ID NO: 114, or SEQ ID

NO: 115.

For example, a polypeptide having the activity of an aminotransferase described herein can have at least 50%, at least 60%, or at least 70%. sequence Identity or homology (e.g., at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%>, at least 98%, at least 99%, or 100%)) to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167- 181. Alternative names for an aminotransferase include, but are not limited to, ω- transaminase, class HI aminotransferase, mcetylormthine aminotransferase, ornithine aminotransferase, omega-amino acid—pymvate aminotransferase, 4-aminobutyrate aminotransferase, DAP A aminotransferase, 2, 2-dialkylglycine decarboxylase, taurine- pyruvate aminotransferase, and glutamate-l-semialdehyde aminotransferase.

It will b appreciated that a number of nucleic acids can encode a polypeptide having a particular amino acid sequence. The degeneracy of the genetic code is well known to the art; i.e., for many amino acids, there is more than one nucleotide triplet that serves as the codon for the amino acid. For example, eodons in the coding sequence for a given enzyme can be modified such that optimal expression in a particular species (e.g., bacteria or fungus) is obtained, using appropriate codon bias tables for that species.

Functional fragments of any of the enzymes described herein can also be used in the methods of the document. The term "functional fragment" as used herein refers to a peptide fragment of a protein that has at least 25% (e.g., at least 30%; at least 40%; at least 50%»; at least 60%; at least 70%; at least 75%; at least 80%; at least 85%; at least 90%; at least 95%; at least 98%; at least 99%; at least 100%; or eve greater than 100%) of the activity of the corresponding mature, full-length, wild-type protein. The functional fragment can generally, but not always, be comprised of a continuous region of the protein, wherein the region has functional activity.

This document also provides (i) functional variants of the enzymes used in the methods of the document and (ii) functional variants of the functional fragments described above. Functional variants of the enzymes and functional fragments can contain additions, deletions, or substitutions relative to the corresponding wild-type sequences. Enzymes with substitutions will generally have not more than 50 (e.g., not more than one, not more than two, not more than three, not more than four, not more than five, not more than six. not more than seven, not more than eight, not more than nine, not more than ten, not more than 12, not more than 15, not more than 20, not more than 25, not more than 30, not more than 35, not more than 40, or not more than 50) amino acid substitutions (e.g., conservative substitutions). This applies to any of the enzymes described herein and functional fragments. A conservative substitution is a substitution of one amino acid for another with similar characteristics, Conservative substitutions include substitutions within the following groups: valine, alanine and glycine; leucine, valine, and isoleucine; aspartic acid and glutamic acid; asparagine and glutamine; serine, cysteine, and threonine; lysine and arginine; and phenylalanine and tyrosine. The nonpolar hydrophobic amino acids include alanine, leucine, isoleucine, valine, proline, phenylalanine, tryptophan and methionine. The polar neutral amino acids include glycine, serine, threonine, cysteine, tyrosine, asparagine and glutamine. The positively charged (basic) amino acids include arginine, lysine and histidine. The negatively charged (acidic) amino acids include aspartic acid and glutamic acid. Any substitution of one member of the above-mentioned polar, basic or acidic groups by another member of the same group can be deemed a conservative substitution, By contrast, a nonconservative substitution is a substitution of one amino acid for another with dissimilar characteristics.

Deletion variants can lack one. two, three, four, five, six, seven, eight, nine, ten, 11. 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acid segments (of two or more amino acids) or non-contiguous single amino acids. Additions (addition variants) include fusion proteins containing: (a) any of the enzymes described herein or a fragment thereof; and (b) internal or terminal (C or N) irrelevant or heterologous amino acid sequences. In the context of such fusion proteins, the term "heterologous amino acid sequences" refers to an amino acid sequence other than (a), A heterologous sequence can be, for example a sequence used for purification of the recombinant protein (e.g., FLAG, polyhistidine (e.g., hexahistidine (SEQ ID NO: 166)), hemagglutinin (HA), ghitathione-S -transferase (GST), or maltosebinding protein ( BP)). Heterologous sequences also can be proteins useful as detectable markers, for example, luciferase, green fluorescent protein (GFP), or chloramphenicol acetyl transferase (CAT), in some embodiments, the fusion protein contains a signal sequence from another protein. In certain host cells (e.g., yeast host cells), expression and/or secretion of the target protein can be increased through use of a heterologous signal sequence, in some embodiments, the fusion protein can contain a carrier (e.g., LH) useful, e.g., in eliciting an immune response for antibody generation) or ER or Goigi apparatus retentio signals. Heterologous sequences can be of varying length and in some cases can be a longer sequences than the full-length target proteins to which the heterologous sequences are attached. Engineered hosts can naturally express none or some (e.g., one or more, two or more, three or more, four or more, five or more, or six or more) of the enzymes of the pathways described herein. Thus, a pathway within an engineered host can include all exogenous enzymes, or can include both endogenous and exogenous enzymes, Endogenous genes of the engineered hosts also can be disrupted to prevent the formation of undesirable metabolites or prevent the loss of intermediates in the pathway through other enzymes acting on such intermediates. Engineered hosts can be referred to as recombinant hosts or recombinant host cells. As described herein, recombinant hosts can include nucleic acids encoding one or more of a methyltransferase, a synthase, β-ketothiolase, a dehydratase, a hydraiase, a dehydrogenase, an. esterase, a thioesterase, a reversible CoA-ligase, a CoA-transferase, a reductase, deacetylase, N~acetyltransferase or an aminotransferase as described in more detail below.

in addition, the production of one or more difunctional products having an odd number of carbon atoms (i.e., C _2t , ₊ 3 building blocks, wherein n is an. integer greater than or equal to one, such as Cs, C ₇ , C9, Cp., C13., Ci$, C37, or (¾ building blocks), can be performed in vitro using the isolated enzymes described herein, using a lysate (e.g., a cell lysate) from a host microorganism as a source of the enzymes, or using a plurality of lysates from different host microorganisms as the source of the enzymes.

Enzymes Generating the C2.1H-3 Aliphatic Backbone for Conversion to Cm _* 3 Building Blocks

As depicted in FIGs. 1-3, a aliphatic backbone H ₃ COC(=0)(CH ₂ )2n ₊ iC(=0)S- ACP or H ₃ COC( ^:::: 0)(CH2)2n- _M C(=0)S-CoA for conversion to one or more Csa+3 building blocks can be formed from propancdioyl~[acp], or acetykCoA and propanedioyl-CoA, via n cycles of methyl-ester shielded carbon chain elongation associated with biotin synthesis using polypeptides having the activity of one or more either NADH or ADPH dependent enzymes, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight.

For example, when n is 1 , the aliphatic backbone is pentanedioyl~[aep] methyl ester or pentanedioyl-CoA methyl ester. For example, when n is 2, the aliphatic backbone is heptanedioyl- acp] methyl ester or heptanedioyl-CoA methyl ester. For example, when n is 3, ihe aliphatic backbone is nonanedioyl-[acp] methyl ester or nonanedioyl-CoA methyl ester. For example, when n is 4, the aliphatic backbone is undecanedi.oyi-[acp] methyl ester or undecanedioyl-CoA methyl ester. For example, when n is 5. the aliphatic backbone is tridecanedioyl~[acp] methyl esier or trideeanedioyl-CoA methyl ester. For example, when n is 6, ihe aliphatic backbone is pentadecanedioyl-[acp] methyl ester or pentadecaaedioyl-CoA methyl ester. For example, when n is 7, the aliphatic backbone is heptadecanedioyl-jacp] methyl ester or heptadecanedioyi-CoA methyl ester. For example, when n is 8, the aliphatic backbone is nonadecanedioyl-[acp] methyl ester or nonadecanedioyl-CoA methyl ester. See Table 1.

In some embodiments, a methyl ester shielded carbon chain elongation associated with biotin biosynthesis route comprises using a polypeptide having the activity of S-adenosyl-L~ methionine (SAM) dependent methyltransferase to form a propanedioyl-[acp] methyl ester, and then performing n cycles of carbon chain elongation, wherein n is an integer greater than or equal to one, such as, for example, one, two. three, four, five, six, seven, or eight, using one or more polypeptides having the activity of one or more of a β-ketoacyl-facp] synthase, a 3- oxo cyl- acpj reductase, a 3-hydroxyacyl-[ cp] dehydratase, and an enoyl-facp] reductase. in some embodiments, a methyl ester shielded carbon chain elongation route comprises using a polypeptide having the activity of S-adenos l-L-methionine (SAM)- dependent methyltransferase to form a propanedioyl-CoA methyl ester, and then performing n cycles of carbon chain elongation, wherein n is an integer greater than or equal to one, such as, for example, one. two, three, four, five, six, seven, or eight, using one or more polypeptides having the activity of one or more of (i) a β-ketothiolase or a β-Κβί:οασ)>1~[αερ] synthase, (ii) an acetoacetyl-CoA reductase, a 3-oxoacyl-[acpj reductase, or a 3- hydroxybutyryl-CoA dehydrogenase, (iii) enoyl-CoA hydratase, and (iv) a trans-2-enoyi-CoA reductase.

In some embodiments, a methyltransferase can be an S-adenosyl-L- ethionine (SAM)- dependent methyltransferase classified, for example, under EC 2.1 , 1.1 7, such as the gene product of hioC, For example, in some embodiments, a polypeptide having the activity of a methyltransferase or S-adenos l-L-methionine (SAM) -dependent methyltramferase is a Bacillus cere s S-adenosyl-L-methionine (SAM)- ependent methyltransferase (see UniProt B Accession No, Q73 I II (SEQ ID NO: 52)). See, for example, Lin, 2012, Biotin Synthesis in Escherichia coli, Ph.D. Dissertation, University of Illinois at Urbana- Champaign).

In some embodiments, a β-ketothiolase may be classified, for example, under EC 2.3, 1.-, such as, for example, EC 2.3.1 .16, such as the gene product of bkiB, such as, for example, a Cupriavid s necator β-keiothiolase (see UniProtKB Accession No. Q0KBP1 (SEQ ID NO: 17)). The β-ketothiolase encoded by bktB from Cupriavidus necator (SEQ ID NO: 17) can accept propanoyl-CoA and pentanedioyl-CoA as substrates, forming a CoA- activated C? aliphatic backbone (see, e.g., Haywood et αί, FEMS Microbiology Letters, 1988, 52:91-96: Slater et ai, J, Bacterial, 1998, 180(8): 1979 - 1987).

in some embodiments, a -ketoacyl~[acp] synthase may be classified, for example, under EC 2.3. I .- (e.g., EC 2.3.1 ,41, EC 2.3.1.179, or EC 2.3, 1.180), such as the gene product of fabB, fabF, or fab H. In some embodiments, a polypeptide having the activity of a β- ketoacyi-facp] synthase is classified under EC 2.3.1.41 , such as the gene product of fabB, such as a Xanthomonas axonopodis pv. citri (strain 306) β-ketoacyl- acp] synthase (see UniProtKB Accession No. Q8PGJ1 (SEQ !D NO: 14)). In some embodiments, a polypeptide having the activity of a fJ~ketoacyI-[acp] synthase is classified under EC 2,3.1.179, such as the gene product of fabF, such as a Xanthomonas axonopodis pv, citri (strain 306) 8-ketoacyl- [acp] synthase (see UniProtKB Accession No, Q8PNE3 (SEQ ID NO: 15)). In some embodiments, a polypeptide having the activity of a β~ΙΐβίοαογΙ~[αορ] synthase is classified under EC 2.3.1.180, such as the gene product of fabH„ such as a Xanthomonas axonopodis pv. citri (strain 306} β-ketoacyl-facp J synthase (see UniProiKB Accession No. Q8PNE8 (SEQ ID NO: 16)).

in some embodiments, a 3-hydroxyacyl-CoA dehydrogenase may be classified, for example, under EC 1.1.1.-, such as, for example, EC 1 .1.1.35, EC 1 , 1.1.36, or EC 1.1.1.157, In some embodiments, a 3~hydroxyacyl-CoA dehydrogenase may be classified under EC 1 ,1.1.35, such as the gene product offadB (e.g., a Staphylococcus aureus 3-hydroxyacyl-CoA dehydrogenase (see UniProtKB Accession No. Q93S 2 (SEQ ID NO: 2)). In some embodiments, a 3-hydroxyacyl-CoA dehydrogenase may be classified under EC 1.1.1.36, such as the gene product of phaB (e.g., Cupriavidus necator aceioacetyl-CoA reductase (see UniProtKB Accession No. P14697 (SEQ ID NO: 3))). A 3-hydroxyacyl-CoA dehydrogenase classified under EC 1.1.1.36 can also be referred to as a acetoacetyl-CoA reductase. See, for example, Liu & Chen, Appi. Microbiol. Biotechnol, 2007, 76(5), 1 153 - 1159; Shen et al , Appl Environ. Microbiol , 201 1, 77(9), 2905 - 2915; or Budde et al, J. Bacteriol, 2010, 192(20), 5319 - 5328, in some embodiments, a. 3~hydroxyacyl~CoA dehydrogenase may be classified under EC 1.1.1.157, such as the gene product of kbd (e.g., a Clostridium acetobutylicum 3-hydroxybutyryl-CoA dehydrogenase (see UniProtKB Accession No. P52041 (SEQ ID NO; 4)). A 3~hydroxyacyl-CoA dehydrogenase classified under EC 1.1.1 , 157 can also be referred to as a 3-hydroxybutyryl-CoA dehydrogenase.

In some embodiments, a 3~oxoacyl-CoA reductase may be classified, for example, under EC 1.1 , 1.100, such as the gene product of fabG (e.g.. an Escherichia coli 3-oxoacyl- [acp] reductase (see UniProtKB Accession No. PGAEK2 (SEQ ID NO: 5)). See, for example, Budde et al , 2010, supra: Nomura et al , Appl Environ. Microbiol, 2005, 71(8), 4297 - 4306).

In some embodiments, an enoyl-CoA hydratase may be classified, for example, under EC 4.2.1 ,17. EC 4.2.1 ,1 19. or EC 4.2.1.150, In some embodiments, an enoyl-CoA hydratase may be classified under EC 4,2.1.17, such as the gene product of crt (e.g., a Clostridium Botulinum enoyl-CoA hydratase (see UniProtKB Accession No. Α5Ϊ6Τ1 (SEQ ID NO: 47)). In some embodiments, an enoyl-CoA hydratase may be classified under EC 4.2.1.1 19, such as the gene product of phaJ (e.g., an Aeromorias caviae enoyl-CoA hydratase (see UniProtKB Accession No. 032472 (SEQ ID NO: 48)) See. for example, Shen et al, 2011, supra', or Fukui et al, J. Bacteriol , 1998, 180(3), 667 - 673. In some embodiments, an enoyl-CoA hydratase may be classified under EC 4.2.1.150. such as the gene product of crt (e.g., a Clostridium acetobutylicum enoyl-CoA hydratase (see UniProtKB Accession No, P52046 (SEQ ID NO: 49)).

In some embodiments, an enoyi-facp] dehydratase such as a 3-hydroxyacyl-facp] dehydratase may be classified, for example, under EC 4.2.1.59, such as the gene product of fabZ (e.g., an Escherichia coli 3~hydroxyacyl-[acp] dehydratase (see UniProtKB Accession No. P0A6Q6 (SEQ ID NO: 1)).

in some embodiments, a trans~2-enoyl~CoA reductase may be classified, for example, under EC 1.3.1.- (e.g., EC 1 ,3.1.8, EC 1.3.1 ,38, or EC 1.3.1.44). such as the gene product of ter (Nishimaki et al , J. Biochem., 1984. 95, 1315 - 1321 ; Shen et al, 201 1 , supra) or tdter (Bond- Watts et al . Biochemistry, 2012, 51. 6827 ^■■■ · 6837). In some embodiments, a trans-2~ enoyl-CoA reductase may be classified under EC 1.3.1.44, such as the gene product of fabV , (e.g., a Treponema denticol trans-2~enoyl-CoA reductase (see UniProtKB Accession No. Q73Q47 (SEQ ID NO; 1 14))) or ter (e.g., an Euglena gracilis (rans-2-enoyl-CoA reductase (see UniProtKB Accession No. Q5EU90 (SEQ ID NO: 1 15))), In some embodiments, a trans-2-enoyl-CoA reductase may be classified under EC 1.3, 1.38, such as the gene product of MSMEG_2155 (e.g., a Mycobacterium smegmatis trans-2-enoyl-CoA reductase (see UniProtKB Accession No. A0QUC2 (SEQ ID NO: 7)).

In some embodiments, an enoyl-[acp] reductase may be classified, for example, under EC 1.3.1.9 or EC 1 .3.1.10. In some embodiments, an enoyl-facp] reductase may be classified. under EC 1.3.1.9, such as the gene product of fabl (e.g., an Escherichia coil enoyl-[aep] reductase (see UniProtKB Accession No. P0AEK4 (SEQ ID NO: 46)). In some embodiments, an enoyl-facp] reductase may be classified under EC 1.3.1.10, such as the gene product of fabl (e.g., a Streptococcus pneumoniae enoyl~[acp] reductase (see UniProtKB Accession No. A0A0X9PXJ6 (SEQ ID NO: 6)).

In some embodiments, an esterase may be classified, for example, under EC 3.1 .1.1.

In some embodiments, an esterase may be classified under EC .3.1 .1.1, such as the gene product oiybflC (e.g., Bacillus subtilis esterase (see UniProtKB Accession No. 031452 (SEQ ID NO: 50)}, hi some embodiments, an esterase may be the gene product of estA from Streptomyces diastatockromogenes (see UniProtKB Accession No. Q59837 (SEQ ID NO: 51)).

Enz mes Generating Terminal Carfooxy! Groups in the Biosynthesis of Cs _n+ s Bmidmg Blocks

As depicted in FIG, 4, a terminal carboxyl group can be enzymatic-ally formed using one or more polypeptides having the activity of one or more of a thioesterase, an aldehyde dehydrogenase, a 7~oxoheptanoate dehydrogenase, a 6-oxohex noate dehydrogenase, a CoA- fransferase, or a reversible CoA-ligase, enzymatieally forming a monomethyl carboxylate

ACP or wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight. In some embodiments, the second terminal carboxyl group leading to the synthesis of a C2fH 3 building block is enzymatically formed by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1.1 ,2, EC 3.1.1.5, EC 3.1.2.-, such as EC 3.1 ,2.14, EC .3.1.2.21 , or EC 3.1.2.27. See, for example, Cantu et ai , Protein Science, 2010, 19, 1281 - 1295; Zhuang et al. Biochemistry, 2008, 47(9), 2789 - 2796; or aggert et ai, I Biol Chem. , 1991 , 266(17), 1 1044 - 1 1050).

In some embodiments, a polypeptide having the activity of a thioesterase classified under EC 3.1.2.- may be selected from: a Spongiibacter sp. IMCC21906 thioesterase (see UniProtKB Accession No. A0A0F7M706 (SEQ ID NO: 58)); an Escherichia coli 06:Hl (strain CFT073 / ATCC 700928 / UP EC) thioesterase (see UniProiKB Accession No. P0ADA2 (SEQ ID NO: 59)); an Escherichia coli thioesterase (see UniProiKB Accession No. P0A8Z3 (SEQ ID NO: 60)); a Congregibacter Htoraiis ΚΤ7Ϊ thioesterase (see UniProtKB Accession No. A4A3N9 (SEQ ID NO: 61 )); a Cuphea kookeriana thioesterase (see UniProtKB Accession No. Q39514 (SEQ ID NO: 62)); an Umhell taria californica thioesterase (see UniProtKB Accession. No. Q41634 (SEQ ID NO: 63)); a Bacillus subtiiis thioesterase (see UniProtKB Accession No, P49851 (SEQ ID NO: 64)): a Bacillus subtiiis thioesterase (see UniProtKB Accession No. Q45061 (SEQ ID NO: 65)); a Bacillus subtiiis thioesterase (see UniProtKB Accession No. P I 4205 (SEQ ID NO: 66)); and a Homo sapiens thioesterase encoded by ACOT13 (see UniProtKB Accession No. Q9NPJ3 (SEQ ID NO: 67)), In some embodiments, a polypeptide having the activity of a thioesterase classified under EC 3.1.2.- may be a Spongiibacter sp. IMCC21906 thioesterase (see UniProtKB Accession No. A0A0F7M706 (SEQ ID NO: 58)) or an Escherichia coli 06: HI (strain CFT073 / ATCC 700928 / UPEC) thioesterase (see UniProtKB Accession No. P0ADA2 (SEQ ID NO: 59)). In some embodiments, a polypeptide having the activity of a thioesterase classified under EC 3.1 ,2.- may be an Escherichia coli thioesterase (see UniProtKB Accession No, P0A8Z3 (SEQ ID NO: 60)) or a Congregibacter litorahs KT71 thioesterase (see UniProtKB Accession No. A4A3N9 (SEQ ID NO: 61 )).

In some embodiments, a polypeptide having the activity of a thioesterase classified under EC 3.1.2.14 may be selected from a Agathobacter recialis DSM 17629 thioesterase (see UniProtKB Accession No, D6E2B1 (SEQ ID NO: 68)): a Lactobacills planta m thioesterase (see UniProtKB Accession No. F9ULU3 (SEQ ID NO: 69)); a Des lfovihrio piezophilus thioester se (see UniProiKB Accession No. M1WJV0 (SEQ ID NO: 70)); and a Streptococcus dysgalactiae thioesterase (see UniProiKB Accession No, C5WH65 (SEQ ID NO: 71)), In some embodiments, a polypeptide having the activity of a ihioesierase classified under EC 3, 1.2.14 may be a Agathohacter rectalis DSM 17629 thioesterase (see UniProiKB Accession No. D6E2B1 (SEQ ID NO: 68)).

In some embodiments, a polypeptide having the activity of a thioesterase may be selected from: a Terrisporob cter othiniensis ihioesierase (see UniProiKB Accession No. A0A0B3WUQ1 (SEQ ID NO: 72)); a Th lassospira xiamenemis M-5 ihioesierase (see UniProiKB Accession No. A0A0B4Y4H4 (SEQ ID NO: 73)); a Cellnlosilyticum kntocellum thioesterase (see UniProiKB Accession No. F2JLT2 (SEQ ID NO: 74)); a Clostridium ihernioceUum ihioesierase (see UniProiKB Accession No. A3DJY9 (SEQ ID NO: 75)); a Thermovirga lienii thioesterase (see UniProiKB Accession No, G7V8P3 (SEQ ID NO: 76)); a Spirochaeta smaragdinae thioesterase (see UniProiKB Accession No. E1.RAP4 (SEQ ID NO: 77)); an Opitutus terrae thioesterase (see UniProiKB Accession No. B1ZXQ1 (SEQ ID NO: 78)): a Thermincola poiens thioesterase (see UniProiKB Accession No. D5XAN2 (SEQ ID NO; 79)); a Clostridium sp. CAG:306 thioesterase (see UniProiKB Accession No, R6Q7V8 (SEQ ID NO: 80)); a Citrobacier rodentium thioesterase (see UniProiKB Accession No, D2TLW8 (SEQ ID NO: 81)); a Vibrio shilonii AK1 thioesterase (see UniProiKB Accession No. A6D1N2 (SEQ ID NO: 82)); a Pseudomonas putida CSVS6 thioesterase (see UniProiKB Accession No, L1 6X0 (SEQ ID NO: 83)); an Alteromonas a stralica thioesterase (see UniProiKB Accession No. A0A075P0Y4 (SEQ ID NO: 84)); a Ferrimorias bale rica ihioesierase (see UniProiKB Accession No. E1SPF5 (SEQ ID NO: 85)); a Marine sediment meiagenome thioesterase (see UniProiKB Accession No, A0A0F9W7B7 (SEQ ID NO: 86)); a Shimwellia blaitae thioesterase (see UniProiKB Accession No. I2BBI6 (SEQ ID NO: 87)); a Clostridium sidjidigenes ihioesierase (see UniProiKB Accession No. A0A084JBW2 (SEQ ID NO: 88)); a Clostridium ceilulolyhcum thioesterase (see UniProiKB Accession No. B8I625 (SEQ ID NO: 89)); a Clostridium argentinense thioesterase (see UniProiKB Accession No, A0A0C1QZB7 (SEQ ID NO: 90)); a Cryptobacteriwn curium ihioesierase (see UniProiKB Accession No. C7ML86 (SEQ ID NO: 91)); a Treponema primitia thioesterase (see UniProiKB Accession No. F5YIQ3 (SEQ ID NO: 92)); a Oceanimonas sp. thioesterase (see UniProiKB Accession No. H2FZ27 (SEQ ID NO: 93)); a Paenibacillus sp. illBB 10380 ihioesterase (see UmProtKB Accession No. A0A0D3V4E9 (SEQ ID NO: 94)); an Aerococcus viridans ATCC 11563 ihioesterase (see UniProtKB Accession No, D4YGM6 (SEQ ID NO: 95)); a Lactococcus raffinol ctis 4877 ihioesterase (see UniProtKB Accession No. Ϊ7ΚΪ30 (SEQ ID NO: 96)); a Catahacier hongkongensis ihioesterase (see UniProtKB Accession No, A0A0M2NEM6 (SEQ ID NO: 97)); a Lactobacillus ruminis SPM0211 ihioesterase (see UniProtKB Accession No. F7R2D3 (SEQ ID NO: 98)); a Clostridium sp. DMHC JO ihioesterase (see UniProtKB Accession No. A0A0L8EW05 (SEQ ID NO: 99)); a Closiridiales bacterium oral (axon 876 sir. F0540 ihioesterase (see UniProtKB Accession No, U2CXE7 (SEQ ID NO: 100)); a Clostridium novyi ihioesterase (see UniProtKB Accession No. A0PXB0 (SEQ ID NO: 101)): a Pepioniphiius harel ACS-l 46-V~Sch2b ihioesterase (see UniProtKB Accession No. E4L0C9 (SEQ ID NO: 102)); a Lactobacillus brevis ihioesterase (see UniProtKB Accession No. Q03SR8 (SEQ ID NO: 103)); a Lactobacillus delbrueckli ihioesterase (see UniProtKB Accession No. WP_01.1678490.1, (SEQ ID NO: 104)); a Clostridium perfringens ihioesterase (see GenBank Accession No. WP_ Q1 1591187.1 (SEQ ID NO: 105)); a Treponema azatonuirici m ihioesterase (see UniProtKB Accession No, F5YA29 (SEQ ID NO: 106)); a Hungaiella hathewayi ihioesterase (see GenBank Accession No. ENY95204.1 (SEQ ID NO: 107)); a Bacillus coagulans ihioesterase (see UniProtKB Accession No. F7Z1.I0 (SEQ ID NO: 108)); a Bdellovibrio bacteriovorus ihioesterase (see UniProtKB Accession No. Q6MKA8 (SEQ ID NO: 109)); a Treponema denticola ihioesterase (see GenBank Accession No. WP_0026885Q0,1 (SEQ ID NO: 110)); a Paenibacillus lactis ihioesterase (see UniProtKB Accession No, G4HNN3 (SEQ ID NO: 1 1 1)); a Cuphea palustris ihioesterase (see UniProtKB Accession No. Q39554 (SEQ ID NO: 1 12)); and a Escherichia coll ihioesterase encoded by yclA (see UniProtKB Accession No. B1LH39 (SEQ ID NO: 113)).

in some embodiments, a polypeptide having the activity of a ihioesterase may be selected from: a Terrisporobacter othimensis ihioesterase (see UniProtKB Accession No. A0A0B3WUQ1 (SEQ ID NO: 72)); a Thalassospira xiamenensls M-5 ihioesterase (see UniProtKB Accession No. A0A0B4Y4H4 (SEQ ID NO: 73)); and a CelMosllytlcum leniocellum ihioesterase (see UniProtKB Accession No. F2JLT2 (SEQ ID NO: 74)).

In some embodiments, a polypeptide having the activity of a ihioesterase may be selected from: a Clostridium- thermocellum ihioesterase (see UniProtKB Accession No. A3DJY9 (SEQ ID NO: 75)); a Thermovirga lienii thioesterase (see UniProtKB Accession No. G7V8P3 (SEQ ID NO: 76)); and a Spirochaet smaragdinae thioesterase (see UniProtKB Accession No. E1RAP4 (SEQ ID NO: 77)).

in some embodiments,, a polypeptide having the activity of a thioesterase may be selected from: an Opi utus terrae thioesterase (see UniProtKB Accession No. B1ZXQ1 (SEQ ID NO: 78)); a Thermmcola patens thioesterase (see UniProtKB Accession No. D5XAN2 (SEQ ID NO: 79)); a Clostridium sp. CAG:3G6 thioesterase (see UniProtKB Accession No. R6Q7V8 (SEQ ID NO: 80)); a Citrobacter rodentium thioesterase (see UniProtKB Accession No. D2TLW8 (SEQ ID NO: 81 )); a Vibrio shilonii AK1 thioesterase (see UniProtKB Accession No. A6D1N2 (SEQ ID NO: 82)); a Pseudomonas putida CSV86 thioesterase (see UniProtKB Accession No. L1 M6X0 (SEQ ID NO: 83)); an Alteromon s ausiralica thioesterase (see UniProtKB Accession No. A0A075P0V4 (SEQ ID NO: 84»; a Ferrimonas balearica thioesterase (see UmProtKB Accession No. E1 SPF5 (SEQ ID NO: 85)); a Marine sediment metagenome thioesterase (see UniProtKB Accession No. A0A0F9W7B7 (SEQ ID NO: 86)); and a Shimweliia hlaitae thioesterase (see UniProtKB Accession No. I2BBI6 (SEQ ID NO: 87)).

In some embodiments, a polypeptide having the activity of a thioesterase may be selected from: a Clostridium sulfidigenes thioesterase (see UniProtKB Accession No. AGA084JBW2 (SEQ ID NO: 88»; a Clostridium cellul lyticum thioesterase (see UniProtKB Accession No. B8I625 (SEQ ID NO: 89)); a Clostridium argeniinense thioesterase (see UniProtKB Accession No. A0A0C1QZB7 (SEQ ID NO: 90»; a Cryptobacierium curium thioesterase (see UniProtKB Accession No, C7ML86 (SEQ ID NO: 91)); a Treponema primitia thioesterase (see UniProtKB Accession No. F5YIQ3 (SEQ ID NO: 92»; a Oce nimonas sp. thioesterase (see UniProtKB Accession No. H2FZ27 (SEQ ID NO: 93»; a Paenibacillus sp. IHBB 10380 thioesterase (see UniProtKB Accession No. A0A0D3V4E9 (SEQ ID NO: 94)); an Aerococc s viridans ATCC 11563 thioesterase (see UniProtKB Accession No. D4YG 6 (SEQ ID NO: 95»; a Lactococcus rafflnolactis 4877 thioesterase (see UniProtKB Accession No. I7KI30 (SEQ ID NO: 96»; a Catahacter hongkongensis thioesterase (see UniProtKB Accession No. A0A0M2NEM6 (SEQ ID NO: 97)); a Lactobacillus ruminis SPM0211 thioesterase (see UniProtKB Accession No. F7R2D3 (SEQ ID NO: 98)); a Clostridium sp, DMHC 10 thioesterase (see UniProtKB Accession No. A0A0L8EW05 (SEQ ID NO; 99)}; a Oostridiaks bacterium oral taxon 876 sir. F0540 thioesterase (see IJniProtKB Accession No. U2CXE7 (SEQ ID NO: 100)); and a Clostridium nov i thioesterase (see UniProtKB Accession No. A0PXB0 (SEQ ID NO: 101)),

In some embodiments, a polypeptide having the activity of a thioesterase may be a Peptoniphilus harei ACS-l46-VSch2b thioesterase (see IJniProtKB Accession No. E4L0C9 (SEQ ID NO: 102)).

in some embodiments, a polypeptide having the activity of a thioesterase may be a Firmicutes bacterium CAG:449 thioesterase (see IJniProtKB Accession No. R.6RDZ9 (SEQ ID NO: 1 82)), Clostridium sp. CAG: 798 thioesterase (see UniProtKB Accession No.

R6XLC3 (SEQ ID NO: ί 83)), [Clostridium] ultunense Esp thioesterase (see UniProtKB Accession No, M1 Z1V0 (SEQ ID NO: 184)), Granulicatella elegans ATCC 700633 thioesterase (see UniProtKB Accession No. D0BKN0 (SEQ ID NO: 185)), Haemophilus influenzae (strain ATCC 51907 / DSM 11121 / KW2Q / Rd) thioesterase (see UniProtKB Accession No. P44886 (SEQ ID NO: 186)), Eggerthella sp, CAG:1427 thioesterase (see UniProiKB Accession No. R5FQ35 (SEQ ID NO: 187)), Vibrio mimicus thioesterase (see UniProtKB Accession No. Q07792 (SEQ ID NO: 188)), Sedimentico!a thiotaurim

thioesterase (see UniProtKB Accession No. A0A0F7JXA5 (SEQ ID NO: 189)), Bacteroides fmegoldn CL09703C10 thioesterase (see UniProtKB Accession No. K5D7V3 (SEQ ID NO: 190)), Pseudoalteromonas sp. SW0106-04 thioesterase (see UniProtKB Accession No.

A0A0M9UHQJ. (SEQ ID NO: 191)), Vibrio mytili thioesterase (see UniProtKB Accession No. A0A0C3EBX5 (SEQ ID NO: 192)), Pseudomonas fluoresces^ thioesterase (see

UniProiKB Accession No. A0A0B7DFD2 (SEQ ID NO: 193)), Pseudomonas siutzeri (strain A1501) thioesterase (see UniProtKB Accession No. A4VL40 (SEQ ID NO: 194)), or

Halobacteriovor x marinus (strain ATCC BAA-682 / DSM 15412 / SJ) (Bacteriovorax marinus) thioesterase (see UniProtKB Accession No. E1WY53 (SEQ ID NO: 195)).

In some embodiments, a terminal formyi group leading to the synthesis of a dicarboxylic acid HOaC CHa iCOaH, wherein n is an integer greater than or equal to one, such as. for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having the activity of an aceylating aldehyde dehydrogenase classified, for example, under EC 1.2.1.10, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutaraie dehydrogenase classified, for example, under EC 1.2, 1.52. In some embodiments, a polypeptide having the activity of an aceylating aldehyde dehydrogenase classified under EC 1.2.1.10 is the gene product of pduP (e.g., a Salmonella iyphimurium aceiylating aldehyde dehydrogenase (see UniProtKB Accession No. H9L4I6 (SEQ ID NO: 18))). in some embodiments, a polypeptide having the activity of an aceylating aldehyde dehydrogenase is the gene product of pduB (e.g., a Salmonella iyphimurium aceiylating aldehyde dehydrogenase (see UniProtKB Accession No. P37449 (SEQ ID NO: 19)). See, for example, Lan ei al, _} 2013, Energy Environ. ScL, 6:2672 - 2681.

In some embodiments, the first terminal carboxyl group leading to the synthesis of a dicarboxylic acid H02C(C¾)2n+] C(½H _S wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzyma!ically formed by a polypeptide having the activity of an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3. See, for example, Guerrillot & Vandecasteele, Eur. J. Biochem., 1977, 81, 185 - 192. In some embodiments, a polypeptide having the activity of an aldehyde dehydrogenase classified under EC 1.2.1.3 may be a Kibdelosporangium sp, aldehyde dehydrogenase (see UniProtKB Accession No. A0A0 3BN67 (SEQ ID NO: 24)).

in some embodiments, the first terminal carboxyl group leading to the synthesis of a dicarboxylic. acid wherein is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatieally formed by a polypeptide having the activity of dehydrogenase classified, for example, under EC 1.2.1.-. In some embodiments, a dehydrogenase may be a 6-oxohexanoate dehydrogenase classified, for example, under EC 1 ,2, 1.- (e.g., EC 1.2.1.63), such as the gene product of chnE (e.g., an Acinetobacter sp. 6-oxohexanoate dehydrogenase (see UniProtKB Accession No. Q9R2F4 (SEQ ID NO: 9)) or a Rhodococcus sp. 6-oxohexanoate dehydrogenase (see UniProtKB Accession No. Q6RXW0 (SEQ iD NO; 10))). In some embodiments, a dehydrogenase may be a 7-oxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.-, such as the gene product of chnE (e.g., an Acinetobacter sp. 7-oxoheptanoate dehydrogenase (see Uni ProtKB Accession No, Q9R2F4 (SEQ ID NO: 12)) or a Rhodococcus sp. 7-oxoheptanoate dehydrogenase (see UniProtKB Accession No. Q6RXW0 (SEQ ID NO: 13))), See, for example, Iwaki ei at. , Appl. Environ. Microbiol. , 1999, 65(1 1), 5158 - 5162; or Lopez-Sanchez et ah , Appl. Environ. Microbiol , 2010, 76(1), 110 - 118. In some embodiments, a dehydrogenase may be a non-acylating NAD-dependent aldehyde dehydrogenase, such as, for example, the gene product of thriG (e.g., a Sphingo onas m crogoliiabida 7-oxoheptanoate dehydrogenase (see UniProtKB Accession No. D9PTN3 (SEQ ID NO: I I))).

In some embodiments, the first terminal carboxyi group leading to the synthesis of dicarboxylic acid wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having the activity of CoA-transferase (e.g., a glutaconaie CoA-transferase) classified, for example, under EC 2,8,3.12, such as the gene product of gctAB (e.g., an Acidaminococcus fermentans glutaconaie Co A -transferase (see UniProtKB Accession No. Q59111 (SEQ ID NO: 40) and UniProtKB Accession No. Q59112 (SEQ ID NO: 41))). See, for example, Buckel et l, 1981, Eur. J. Biocke ,, 1 18:315 - 321.

In some embodiments, the first terminal carboxyi group leading to the synthesis of a dicarboxylic acid H02C(CH2)2n ₊ iC02H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having th activity of reversible CoA-ligase (e.g., a succinate-CoA ligase) classified, for example, under EC 6,2.1 ,5, such as a Thermococcus kodakaraensis succinate- CoA-ligase. See, for example, Shikata ei al , 2007, J Biol Chem^ 2S2(3?):26963 - 26970. in some embodiments, the second terminal carboxyi group leading to the synthesis of a dicarboxylic acid H02C(CH )2n÷iC02H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having the activity of an esterase (e.g., a carboxylesterase) classified, for example, under EC 3. L1.1 . See, for example, Kunst ei al , 1997, Nature, 390 (6657), 249 - 256. In some embodiments, a polypeptide having the activity of an esterase classified under EC 3.1.1 , 1 may be the gene product of yb/K (e.g., a Bacillus subtilis esterase (see UniProtKB Accession No. 031452 (SEQ ID NO: 50))). In some embodiments, a polypeptide having the activity of an esterase may be the gene product of estA. (e.g., a Sirepiomyces diastatochromogenes esterase (see UniProtKB Accession No, Q59837 (SEQ ID NO: 51)). Enzymes Generating Terminal Amine Gronps in the Biosynthesis of €2 +3 Building Blocks

As depicted in FIGs, 3 and 4. terminal amine groups can be enzymatically formed using a polypeptide having the activity of an aminotransferase or a deacetylase.

In some embodiments, a terminal amine group leading to the synthesis of 7- aminoheptanoic acid is enzymatically formed by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1.-, such as EC 2.6.1.11, EC 2,6, 1.13, EC 2.6.1.18, EC 2.6,1.19, EC 2,6.1.29, EC 2.6,1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4,3.8, such as, for example, the amino acid sequences of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181. Some of these aminotransferases are diamine ω-transaminases (e.g., an Escherichia coli aminotransferase (see UniProtKB Accession No. P42588 (SEQ ID NO: 1 19)). For example, the aminotransferases classified, for example, under EC 2.6.1.29 or EC 2,6.1.82 ma be diamine ω-transaminases.

The reversible aminotransferase from Chrontob cterium violaceum (see UniProtKB Accession No, Q7NWG4 (SEQ ID NO: 116)) has demonstrated analogous activity accepting 6-aminohexanoie acid as amino donor, thus forming the first terminal amine group in adipate semialdehyde (Kaulmann et al.. Enzyme and Microbial Technology, 2007, 41 , 628 - 637).

The reversible 4-aminobubyrate:2-oxoglutarate transaminase from Streptomyces griseus has demonstrated analogous activity for the conversion of 6-axmnohexanoate to adipate semialdehyde (Yonaha e al . Enr. J. Biochem. , 1985, 146: 101 - 106).

The reversible 5~aminovalerate transaminase from Clostridium viride has demonstrated analogous activity for the conversion of 6-aminohexanoate to adipate semialdehyde (Barker et aL 1 Biol Client , 1987, 262(19), 8994 - 9003).

in some embodiments, a terminal amine group leading to the synthesis of an aminocarboxylate H2 (C¾)2n+2C02H or a diamine H2N(CH2) n+3N¾, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having the activity of a diamine ω- trans minase. For example, the second terminal amine group can be enzymaiieally formed by a polypeptide having the activity of a diamine ω-transaminase classified, for example. under EC 2.6.1 .29 or classified, for example, under EC 2.6.1.82, such as the gene product of ygj ^' G from E. coli (e.g., an Escherichia coli aminotransferase (se UniProtKB Accession No. P42588 (SEQ ID NO: 1 19)).

The gene product of ygj ^' G accepts a broad range of diamine carbon chain length substrates, such as putrescine, eadaverine, and spermidine (see, for example, Samsonova et al. , BMC Microbiology, 2003 , 3:2).

The diamine ω-transaminase from E.coli strain B has demonstrated activity for 1,7 diaminoheptane (Kim, The Journal of Chemistry, 1964, 239(3), 783 - 786).

In some embodiments, an aminotransferase classified under EC 2.6, 1.- may be a Chromobacterium violaceum aminotransferase (see UniProtKB Accession No. Q7NWG4 (SEQ ID NO: 116)), a Thermotnicrobium roseum aminotransferase (see UniProtKB Accession No, B9L0N2 (SEQ ID NO: 117» or a Nautella italiea aminotransferase (see UniProtKB Accession No. A0A0H5D6A2 (SEQ ID NO: 1 18)).

In some embodiments, an aminotransferase may be selected from: a Kiloniella spongiae aminotransferase (see UniProtKB Accession No. A0A0H2MDD9 (SEQ ID NO: 120)); a Haematobacter missouriensis aminotransferase (see UniProtKB Accession No. A0A086YIZ0 (SEQ ID NO: 121)); a Mesorhizobium sp, LCI 03 aminotransferase (see UniProtKB Accession No, A0A0H1AH98 (SEQ ID NO: 122)); a Pseudomonas sp. 10-1 B aminotransferase (see UniProtKB Accession No. A0A0E9ZHQ3 (SEQ ID NO: 123)); a Mesorhizobiu alhagi aminotransferase (see UniProtKB Accession No, H0I025 (SEQ ID NO: 124)); a Pseudomonas aeruginosa aminotransferase (see UniProtKB Accession No. Q9HV04 (SEQ ID NO: 125)); a Pseudomonas syringae aminotransferase (see UniProtKB Accession No. Q4ZES9 (SEQ ID NO: 126)); a Rhodobacter sphaeroides aminotransferase (see UniProtKB Accession No. Q3IWE9 (SEQ ID NO: 127)); a Vibrio fluviaiis aminotransferase (see UniProtKB Accession No. F2XBU9 (SEQ ID NO: 128»; a Lutibaculum haratangense AMVl aminotransferase (see UniProtKB Accession No, V4RM39 (SEQ ID NO: 129)); a Truepera radiovictrix aminotransferase (see UniProtKB Accession No. D7CVJ6 (SEQ ID NO: 130)); an Aquamicrobium defluvii aminotransferase (see UniProtKB Accession No, A0A011UWB9 (SEQ ID NO: 131»; a Neorhizobium galegae bv. orientalis str. HA MB I 540 aminotransferase (see UniProtKB Accession No. A0A0683UV9 (SEQ ID NO: 132)); a Microvirga sp, BSC 39 aminotransferase (see UniProtKB Accession No. A0A086MKC4 (SEQ ID NO: 133)); a Mesorhizobium sp. LCI 03 aminotransferase (see UniProtKB Accession No. A0A0H1A7R9 (SEQ ID NO: 134)); a Starkeya novella aminotransferase (see UniProtKB Accession No. D7A1Z2 (SEQ ID NO: 135}); an Azospirillum Hpoferum aminotransferase (see UniProtKB Accession No. G7Z3P2 (SEQ ID NO: 136)); a Variovorax sp. CF313 aminotransferase (see UniProtKB Accession No. J2TM48 (SEQ ID NO: 137); and a Thalassospira profimdimaris WP0211 aminotransferase (see UniProtKB Accession No. K2KXB1 (SEQ ID NO: 138)).

In some embodiments, an aminotransferase may be selected from: a Kiloniella spongiae aminotransferase (see UniProtKB Accession No. A0A0H2MDD9 (SEQ ID NO: 120)); a Haematobacter misso riensis aminotransferase (see UniProtKB Accession No. A0A086YIZ0 (SEQ ID NO: 121)); and a Mesorhizobium sp. LCI 03 aminotransjerase (see UniProtKB Accession No. A0A0H1AH98 (SEQ ID NO: 122)).

In some embodiments, an aminotransjerase may be selected from: a Pseudomonas sp. 10-1 B aminotransferase (see UniProtKB Accession No. A0A0E9ZHQ3 (SEQ ID NO: 123)); a Mesorhizobium alhagi aminotransferase (see UniProtKB Accession No. HO ^' 1025 (SEQ ID NO: 124); a Ther omicrobium roseum aminotransferase (see UniProtKB Accession No. B9L-0N2 (SEQ ID NO: 1 17)); and a Chromobacterium violaceum aminotransferase (see UniProtKB Accession No. Q7NWG4 (SEQ ID NO: 116)).

In some embodiments, an aminotransferase may be selected from a: Vibrio fluvialis aminotransferase (see UniProtKB Accession No, F2XBU9 (SEQ ID NO: 167)); Rhodospirillum centenwn (strain ATCC 51521 / SW) aminotransferase (see UniProtKB Accession No. B61SI5 (SEQ ID NO: 1 8)); marine sediment metagenome aminotransferase (see UniProtKB Accession No, A0A0F9UFF8 (SEQ ID NO: 169)); Acidimicrobium ferrooxidans (strain DSM 10331 / JCM 15462 / 'BRC 103882 / ICP) aminotransferase (see UniProtKB Accession No, C7LZG4 (SEQ ID NO: 170)); Tistrella mobilis (strain KA081Q2Q 065) aminotransferase (see UniProtKB Accession No. I3TH77 (SEQ ID NO: 171)); Pseudomonas iaiwanensis SJ9 aminotransferase (see UniProtKB Accession No. V7D492 (SEQ ID NO: 172)): Methylobacterium aquaticum aminotransferase (see UniProtKB Accession No, A0A0C6G014 (SEQ ID NO: 173)); Candida tenuis (strain ATCC 10573 / BCRC 21748 / CBS 615 / JCM 9827 / NBR.C 10315 / NR.RL Y-1498 / VKM Y-70) (Yeast) aminotransferase (see UniProtKB Accession No, G3BAK1 (SEQ ID NO: 174)); Tepidicaulis marinus aminotransferase (see UniProtKB Accession No. A0A081B6K8 (SEQ ID NO: 175)); Firmicutes bacterium CAG;24 aminotransferase (see UniProiKB Accession No. R5HDC3 (SEQ ID NO: 176)); Pseudooceanicola batsemis (strain ATCC BAA-863 / DSM 15984 / KCTC 12145 / HTCC2597) (Oceanicola batsensis) aminotransferase (see Uni ProtKB Accession No. A3U3W9 (SEQ ID NO: 177)); Defiuviimonas sp. 20V 17 aminotransferase (see UniProtKB Accession No. A0A059IS31 (SEQ ID NO: 178)); Sph gobacterium spiritivorum ATCC 33S61 aminotransferase (see UniProtKB Accession ^' No. D7VKX2 (SEQ ID NO: 179)); Ramlibacter iataouinemis (strain ATCC BAA-407 / DSM 14655 / IMG 21543 / TTB310) aminotransferase (see UniProiKB Accession No. F5Y1J0 (SEQ ID NO: 1 80»; and Bradyrhizobi m sp, DOA9 aminotransferase (see UniProtKB Accession No. A0A0 1M4Q7 (SEQ ID NO: 181)).

In some embodiments, the second terminal amine group leading to the synthesis of a diamine H2N(CH2)2n+3N¾, wherein » is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is enzymatically formed by a polypeptide having the activity of deacetylase such as acetylpntrescine deacetylase classified, for example, under EC 3.5.1.62, in some embodiments, an acetylpuirescine deacetylase classified under EC 3.5.1 ,62 may be the gene product of aphA (e.g., a Burkholderia pseudomallei acetylpuirescine deacetylase (see UniProtKB Accession No. Q3JUN4 (SEQ ID NO: 42), a Pse domonas aeruginosa acetylpuirescine deacetylase (see UniProtKB Accession No. Q9DT5 (SEQ ID NO: 43), or a Mycoplana ramose acetylpuirescine deacetylase (see UniProtKB Accession No. Q48935 (SEQ ID NO: 44)) or aphB (e.g., Pseudomonas aeruginosa acetylpuirescine deacetylase (see UniProtKB Accession No. Q9I6H0 (SEQ ID NO: 45)).

The acetylpuirescine deacetylase from Micrococcus iuteus K~l l accepts a broad range of carbon chain length substrates, such as aeetylputrescine, aeetylcadaverhie and N ⁸ .. aeetylspermidine (see, for example, Suzuki et ai, 1986, BBA - General Subjects, 882( 1 ):140 - 142).

Enzymes Generating the Terminal Hydroxy! Groups in the Biosynthesis of C ^! 2 _!5 +3 Bnildirig Blocks

As depicted in FlGs. 7 and 8. a terminal hydroxy! group can be enzymatically formed using a polypeptide having the activity of an alcohol dehydrogenase, In some embodiments, a terminal hydroxy! group leading to the synthesis of a QH(€H2)2ri-i-iOH is enxymaticaily formed by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., 1, 2, 21, 61, 184, or 258). In some embodiments, a polypepiide having the activity of an alcohol dehydrogenase classified under EC 1.1.1.- is the gene product oiyqhD (e.g., an Escherichia coll alcohol dehydrogenase (see UniProtKB Accession No. Q46856 (SEQ ID NO: 20))) or cpnD (e.g., Comamonas sp.alcohol dehydrogenase (see UniProtKB Accession No. QSGAW4 (SEQ ID NO: 21))). See, for example, Iwaki et al _t 2002, Appl. Environ. Microbiol, 68(1 1 ):5671 - 5684. In some embodiments, a polypeptide having the activity of an alcohol dehydrogenase classified under EC 1 , 1.1 ,2 is the gene product of ADH6 (e.g., Saccharomyces cerevisiae alcohol dehydrogenase (see UniProtKB Accession No. Q04894 (SEQ ID NO: 22))), See, for example, Larroy et ah , 2002, Bloche J, 361(Pt 1), 163 - 172). Irs some embodiments, a polypepiide having the activity of an alcohol dehydrogenase classified under EC 1 , 1 .1.61 is the gene product of gbd (e.g., Cuprividus necator alcohol dehydrogenase (see UniProtKB Accession No. Q59104 (SEQ ID NO; 23))). An alcohol dehydrogenase classified under EC L I .1.61 may also be referred to as a 4-hydroxyhutyrate dehydrogenase. In some embodiments, a polypeptide having the activity of an alcohol dehydrogenase classified under EC 1.1.1.258 is the gene product of chnD (e.g., an Acinetobacter sp, 6-hydroxyhexanoate dehydrogenase (see UniProtKB Accession No. Q7WVD0 (SEQ ID NO: 8))). See, for example, Iwaki et at, Appl. Environ. Microbiol , 1999, supra. An alcohol dehydrogenase classified under EC 1 .1.1 ,258 may also be referred to as a 6-hydroxyhexanoate dehydrogenase.

Other Enzymes Used in the Biosynthesis of Building Blocks

in some embodiments, a polypeptide having the activity of a carboxylate reductase is classified, for example, under EC 1.2.99.6. In some embodiments, a polypeptide having the activity of a carboxylate reductase may be the gene product of car (e.g., a Mycobacterium marinum carboxylate reductase (see UniProtKB Accession No. B2HN69 (SEQ ID NO: 25) or a Nocardia iowemis carboxylate reductase (see UniProtKB Accession No. Q6RKB1 (SEQ ID NO: 26 ^' ))). In some embodiments, a polypeptide having the activity of carboxylate reductase is the gene product ifadD9 (e.g., Mycobacterium smegmatis fatty-acid-CoA ligase (see UniProtKB Accession No. A0QWT7 (SEQ ID NO: 27)) or Mycobacterium smegmatis faity-acid-CoA Ugase (see UniProtKB Accession No. A0A0D6J1A6 (SEQ ID NO: 28))).

In some embodiments;, a carhoxylate reductase may be selected from: a Mycobacterium smegmatis carhoxylate reductase (see UniProtKB Accession No, A0R484 (SEQ ID NO; 29)); a. Mycobacterium avium carhoxylate reductase (see GenBank Accession No. WPJ) 19730046.1 (SEQ ID NO: 30)); a Segnilipar s rugosus carhoxylate reductase (see UniProtKB Accession No. E5XUS9 (SEQ ID NO: 31 )); a Mycobacterium sp, JS623 carhoxylate reductase (see UniProtKB Accession No. L01YJ8 (SEQ ID NO; 32)); a Mycobacterium heckeshornense carhoxylate reductase (see UniProtKB Accession No. A0A0J8.X8T4 (SEQ ID NO: 33)}; a Mycobacterium goodii carhoxylate reductase (see UniProtKB Accession No. A0A0K0XCM7 (SEQ ID NO: 34)); a Mycobacterium goodii carhoxylate reductase (see UniProtKB Accession No. A0A0K0X557 (SEQ ID NO: 35)); a Mycobacterium intracellulare carhoxylate reductase (see UniProtKB Accession No. H8ITF4 (SEQ ID NO: 36)); a Mycobacterium massiliense carhoxylate reductase (see Genbank Accession No. EI VI 1 143.1 (SEQ ID NO: 37)); a Segniliparus rotundas carhoxylate reductase (see Genbank Accession No, D6Z860 (SEQ ID NO: 38)): and a Segniliparus rotundus carhoxylate reductase (see UniProtKB Accession No. D6ZDT1 (SEQ ID NO: 39)).

In some embodiments, a carhoxylate reductase may be selected from; a Mycobacterium smegmatis carhoxylate reductase (see UniProtKB Accession No. A0R484 (SEQ ID NO: 29)); a Mycobacterium avium carhoxylate reductase (see GenBank Accession No. WPJ) 19730046.1 (SEQ ID NO: 30)); and a Segniliparus rugosus carhoxylate reductase (see UniProtKB Accession No. E5XUS9 (SEQ ID NO: 31)),

In some embodiments, a carhoxylate reductase may be selected from: a Mycobacterium sp. JS623 carhoxylate reductase (see UniProtKB Accession No. L0IYJ (SEQ ID NO: 32)): a Mycobacterium heckeshornense carhoxylate reductase (see UniProtKB Accession No. A0A0J8X8T4 (SEQ ID NO: 33)); a Mycobacterium goodii carhoxylate reductase (see UniProtKB Accession No. A0A0 0XCM7 (SEQ ID NO: 34)); a Mycobacterium goodii carhoxylate reductase (see UniProtKB Accession No. A0AQK0X557 (SEQ ID NO: 35)); a Mycobacterium intracellulare carhoxylate reductase (see UniProtKB Accession No, H8ITF4 (SEQ ID NO: 36)); and a Mycobacterium smegmatis fatty-acid-CoA Ugase (see UniProtKB Accession No. A0A0D6J1 A6 (SEQ ID NO: 28)). In some embodiment, a carboxylate reductase may be selected from a: Chromera velia CCMP287S carboxylate reductase (see UniProtKB Accession No. A0A0G4ID64 (SEQ ID NO: 196)); Cyberlindnera j dinii (Ferula yeast) (Pichia jadinii) carboxylate reductase (see UniProtKB Accession No. A0A0H5CAG1 (SEQ ID NO: 197)); Pest lotiopsis fid W106- 1 carboxylate reductase (see UniProtKB Accession No. W3XHR4 (SEQ ID NO: 198)); Caenorhabditis elegans carboxylate reductase (see UniProtKB Accession No. Q 18660 (SEQ ID NO: 199)); Tetrahymena thermophila (strain SB210) carboxylate reductase (see UniProtKB Accession No. I7MB41 (SEQ ID NO: 200)); Auxenochlorella protolhecoides (Green microalga) (Chlorella protoihecoides) carboxylate reductase (see UniProtKB Accession No. A0A087SHC7 (SEQ ID NO: 201)); Lichtheimia corymbifera JMRC:FSU:9682 carboxylate reductase (see UniProtKB Accession No. A0A068SDQ8 (SEQ ID NO: 202)); Labilithrix luteola carboxylate reductase (see UniProtKB Accession No, A0A0K1PNT5 (SEQ ID NO: 203)); Geolrichum candidu (Oospora lactis) (Dipodascus geoirlckum) carboxylate reductase (see UniProtKB Accession No, A0A0J9XGX9 (SEQ ID NO: 204)); Kuraishia capsulata CBS 1993 carboxylate reductase (see UniProtKB Accession No. W6MHS7 (SEQ ID NO: 205)); Phytophthora soj e (strain P6497) (Soybean stem and root rot agent) (Phytophthora megasperma f. sp, glycines) carboxylate reductase (see UniProtKB Accession No. G4YTV4 (SEQ ID NO: 206)); Dictyosteliu discoideum (Slime mold) carboxylate reductase (see UniProtKB Accession No. Q1ZXQ4 (SEQ ID NO: 207)); Ascaris suum (Pig roundworm) (Ascaris lumbricoides) carboxylate reductase (see UniProtKB Accession No. FI XIL (SEQ ID NO: 208)); Nocardia brasiliensis NBRC 14402 carboxylate reductase (see UniProtKB Accession No. A0A034UK40 (SEQ ID NO: 209)): Rhizopus microspores carboxylate reductase (see UniProtKB Accession No. A0A0C7BIS0 (SEQ ID NO: 210)); Theileria parva (East coast fever infection agent) carboxylate reductase (see UniProtKB Accession No. Q4N8F1 (SEQ ID NO: 211)); Anisakis simplex (Herring worm) carboxylate reductase (see UniProtKB Accession No. A0A0M3J210 (SEQ ID NO: 2 2)); Helobdella robusta (Califomian leech) carboxylate reductase (see UniProtKB Accession No. T1EG09 (SEQ ID NO: 213)); Mycobacterium lepromatosis carboxylate reductase (see UniProtKB Accession No, A0A0F4ES51 (SEQ ID NO: 214)); and Schizopora paradoxa carboxylate reductase (see UniProtKB Accession No. A0A0H2RRC5 (SEQ ID NO: 215)). In some embodiments, a polypeptide having the activity of an N-aeeiyhransferase (e.g., a lysine N-acetyltransferase) is classified, for example, under EC 2.3.1.32. See, for example, Paik et al , Arch Biochem Biophys., 1964 Nov., 108: 221-29. An N~ cetyliransferase classified under EC 2.3.1.32 may be the gene product of LYC1 (e.g., Yarwwia Hpolytlca lysine N-acetyltransferase (see UniProtKB Accession No. P41929 (SEQ ID NO: 53))).

In some embodiments, a polypeptide having the activity of a phosphopantetheine transferase is classified, for example, under EC 2.7.8.-, such as EC 2.7,8.7. in some embodiments, a phosphopantetheine transferase classified under EC 2.7.8.- is the gene product of sfp (e.g., Bacillus suhtilis phosphopantetheine transferase (see UniProtKB Accession No. P39135 (SEQ ID NO: 54)). In some embodiments, a phosphopantetheine transferase classified under EC 2,7.8.7 is the gene product of npt (e.g., Nocardia sp. NRRL 5646 phosphopantetheine transferase (see Genbank Accession No. ABI83656.1 (SEQ ID NO: 55))). In some embodiments, a phosphopantetheine transferase may be the gene products of griC and griD from Streptomyces griseus (UniProtKB Accession No, Q9ZN75 (SEQ ID NO: 56) and UniProtKB Accession No. Q9ZN74 (SEQ ID NO: 57)). See, for example, Suzuki et al., J. Aniihiot., 2007, 60(6), 380 - 387.

Eraymes Attenuated to Improve Biosynthesis of C . _? Building Blocks

One or more of the following enzymes may be attenuated in a recombinant host described herein, such as, for example, a polyhydroxyalkanoate synthase, an acetyl-CoA thioesterase, an acetyl-CoA specific β-ketothiolase, a phosphotransacetylase forming acetate, an acetate kinase, a lactate dehydrogenase, a menaquinol-fumarate oxidoreductase, a 2- oxoacid decarboxylase producing isobutanol, an alcohol dehydrogenase forming ethanol, a triose phosphate isomerase, a pyruvate decarboxylase, a glucose -6-phosphate isomerase, a transhydrogenase dissipating the NADH or NADPH imbalance, an glutamate dehydrogenase dissipating the NADH or NADPH imbalance, a NADH/NADPH-utilizing glutamate dehydrogenase, a pimeloy -CoA dehydrogenase; an acyl-CoA. dehydrogenase accepting C _? , C ₇ , (¾, C;i, C| 3, Cis, C{7, or C _f y building blocks and central precursors as substrates; a glutaryl-CoA dehydrogenase: or a pimeloyl-CoA synthetase. In some embodiments, a polyhydroxyalkanoate synthase is attenuated, such as the gene product of phaC, from, for example Rhodospirilium riihr um (see IJnlProtKB Accession No. Q9 NU7 (SEQ ID NO: 141)), In some embodiments, a lactate dehydrogenase is attenuated, such as the gene product of IdhA (Shen et ah, 2011, supra) or Idh, such as, for example, the gene product of idh from Geobacillus siearothermophilus (see UniProtKB Accession No. P00344 (SEQ ID NO: 142)). In some embodiments, a menaquinol-fumarate. oxidoreductase, such as the gene product of frdBC (see, e.g., Shen et ai. , 201 1 , supra), is attenuated.

In some embodiments, a triose phosphate isomerase classified, for example, under EC 5.3,1 , 1, is attenuated, such as, for example, the gene product of tpiA from, for example, Escherichia coli (see UniProtKB Accession No. P0A858 (SEQ ID NO: 143)). In some embodiments, a glucose-6-phosphate isomerase classified, for example, under EC 5.3.1 ,9, such as the gene product of GPI from, for example. Homo sapiens (see UniProtKB Accession No, P06744 (SEQ ID NO: 144)) is attenuated. In some embodiments, a glutamate dehydrogenase dissipating the NADH or NADPH imbalance classified, for example, under EC 1.4.1.2 (NADH-specific), EC L4.1.3, or EC 1 ,4.1.4 (NADPH-specific) is attenuated. In some embodiments, an NADH/NADPH-utilizing glutamate dehydrogenase classified, for example, under EC 1 ,4.1 ,3 is attenuated, such as, for example an NADH NADPH-utilizing glutamate dehydrogenase from Homo sapiens (see UniProtKB Accession No. P00367 (SEQ ID NO: 145)) or Bos Taurus (see UniProtKB Accession No, P00366 (SEQ ID NO: 146)),

In some embodiments, a pimeloyl-CoA dehydrogenase classified, for example, under EC 1 ,3.1.62, such as the gene product oiphnD from, for example, Aromatoleum aromaiicum (see UniProtKB Accession No. Q5P017 (SEQ ID NO: 147)) is attenuated, in some embodiments, an acyl-CoA. dehydrogenase accepting C5, C7, C9, Cj i, C13, C15, C17, or C1 building blocks and central precursors as substrates classified, for example, under EC 1.3,8.1 , EC 1.3.8.7, EC 1 .3.8.8, or EC 1.3,8,9 is attenuated, such as, for example, an acyl-CoA dehydrogenase from Rattus norvegicus (see UniProtKB Accession No, PQ8503 (SEQ ID NO: 148)),

In some embodiments, a glutaryl-CoA dehydrogenase classified, for example, under EC 1.3.8.6 is attenuated, such as a glutaryl-CoA dehydrogenase from Kibdelosporangium sp. MJ126-NF4 (see UniProtKB Accession No. A0A0K3B4X3 (SEQ ID NO: 149)), In some embodiments, a pimeloyl-CoA synthetase classified, for example, under EC 6,2.1.14 is attenuated, such as the gene product of BIOl from, for example, Saccharomyces cerevisiae (see UniProtKB Accession No. E9P9F6 (SEQ ID NO: 150)).

Generic Biochemical Pathways

The generic biochemical pathways described herein are illustrated in FIGs. 1.-8. In all figures, n is an integer greater than or equal to one, such as, for example, one, two. three, four, five, six, seven, or eight. The integer n corresponds to the number of methyl ester shielded carbon chain elongation cycles used to synthesize an aliphatic carbon chain backbone having an odd number of carbon atoms from (i) acetyl-CoA and propanedioyl-CoA or (ii) propanedioyl-[acp]. For example, the aliphatic carbon chain backbone resulting from n cycles of methyl ester shielded carbon chain elongation will have (2n+3) carbon atoms. See Table 1 . Table 1 lists the€21-1+3 aliphatic carbon chain backbones produced using the methods below after n cycles of methyl ester shielded carbon chain elongation for n is 1, 2, 3, 4, 5, 6, 7, or 8. Chemical structures for the Can+3 aliphatic carbon chain backbones (i.e., carboxyl- [acp] methyl esters and carboxyl-CoA methyl esters) of varying carbon chain length are illustrated in FIGs. 9 and 10.

Table 1, C _2n+ 3 Aliphatic Backbones

norsadecanedioyi-CoA methyl ester

Tables 2A, 2B, and 2C list intermediate compounds synthesized in the production of £2*1+3 building blocks from a C&n-s aliphatic backbone produced from (i) acetyl-CoA and propanedioyl-CoA via n cycles of methyl ester shielded carbon chain elongation or (ii) propanedioyl-[acp] via n cycles of methyl ester shielded carbon chain elongation, wherein n is 1 , 2, 3, 4, 5, 6, 7, or 8. The chemical structures for intermediate compounds of varying carbon chain lengths are illustrated in FIGs. 1 1 (monomethyl earboxylates), 12 (monomethyi carboxylate semialdehdyes), 14 (monomethyl ammoearboxylates), 17 (aminoaldeh des), 20 (hydroxyamines), 21 (acetamidocarboxylates), 22, (aeetarmdoaldehydes), 23 (aceiamidoamines), 24 (dials), 25 (hydroxyaldehydes).

In addition, FIGs, 27 (3-oxo-carboxyl-ACP methyl esters), 28 (3-oxo-carboxyl-CoA methyl esters), 29 (3-hydroxy-carboxyl-ACP methyl esters), 30 (3-hydroxy-carboxyl-CoA methyl esters), 31 (2,3-dehydrocarboxyl-ACP methyl esters), and 32 (2,3-dehydrocarhoxyl- ACP methyl esters) illustrate the structures of intermediates produced during methyl ester shielded carbon chain elongation cycles.

Tables 3A and 3B list building blocks, which are difunctional products having an odd number of carbon atoms, synthesized by the enzymatic conversion of a Ο η+3 aliphatic backbone produced from (i) aceiyl-CoA and propanedioyl-CoA via n cycles of methyl ester shielded carbon chain elongation or (ii) propanedioyl- [acp J via n cycles of methyl ester shielded carbon chain elongation, wherein n is 1, 2, 3, 4, 5, 6, 7, or 8. The chemical structures for C211+3 building blocks of varying carbon chain length (e.g., C5, C ₇ , C¾, Cn, Co, C\$, Ct? _> and C 39 building blocks) are illustrated in FiGs. 13 (dicarboxylic acids). 15 (ammoearboxylates), 16 (carboxylate semialdehydes), 18 (diamines), 19 (hydroxycarboxylates), and 26 (diols).

Table 2C. Intermediates m the Prodfuetjoss i>f C _2a+3 BoiMiis Blocks (coni.)

Table 3A. C _2lrf Bmidlag Blocks

Table 3 , Cj _B . _f 3 Building Blocks (com

HO(CH ₂ ) _2!1+ jC0 ₂ e MO(CH ₂ ) _2il+3 OH

Pathways Using NADPH-Specifk Enzymes to Produ e a Carboxyl-ACP Methyl Ester as a Central Precursor Leading to Difasietiosia! Products

In some embodiments, a C _2n*3 aliphatic backbone H ₃ COC( >)(CH2)2 _fl ÷iC( ))S-ACP, also referred to as a carboxyi-ACP methyl ester, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central metabolite propanedioyl-[acpj. First, propanedioyl-f acp] is converted to propanedioyl-[acp] methyl ester by a polypeptide having the activity of a S~adenosyl-L~ methionine (SAM) -dependent methyltrcmsferase classified, for example, under EC 2.1.1.197 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52). Second, propanedioyl-[acp] methyl ester is enzymatieaiiy converted to a carboxyl-ACP methyl ester via n cycles of methyl-ester shielded carbon chain elongation, wherein n is an integer greater Chan or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight. See FIG. 1. Each cycle m of the n cycles of methyl-ester shielded carbon chain elongation includes: the conversion of H ₃ COC(==0)(CH ₂ )2m-iC( ^::: 0)S-ACP with propanedioyl-[acp] to a 3-oxo-earboxyj-ACP methyl ester I-!. ₃ COC(=0)(CH2)2m-iC(=0)CH ₂ C(-0)S-ACP by a polypeptide having the activity of a synthase classified, for example, under EC 2,3.1.- (e.g., EC 2.3.1 ,41 or EC 23.1.179) when m is greater than 1 or EC 2.3.1.41, EC 2.3, 1 .179, or EC 2.3,1.180 when m is 1 (i.e., during the first cycle of methyl-ester shielded carbon chain elongation)) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to a 3-hydroxy~carboxyl-ACP methyl ester I-l3COC(=0)(C¾)2 _m . iCH(OH)CH ₂ C( ^: =0)S~ACP by a polypeptide having the activity of a 3-oxoa yl-[acpJ reductase classified, for example, under EC 1.1 ,1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to a 2,3-dehydrocarboxyl~ACP methyl ester e ₃ COC(=0)(CH ₂ ) ₂ m-jCH=CHC(-0)S-ACP by a polypeptide having the activity of a 3- hydroxyacyi-facpj dehydratase classified, for example, under EC 4,2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 1 ); followed by conversion to H ₃ COC( ^::: O)(C¾)2 _m +iC(=0)S-ACP by a polypeptide having the activity of an enoyl-f cp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6). See FIG. 1.

Pathways Usirag NADFH-Specific Enzymes to Produce a Carbox l-CoA Methyl Ester as a Central Precursor Leading to Difimctienal Products

In some embodiments, a C2n+3 aliphatic backbone

also referred to as a carboxyl-CoA methyl ester, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central metabolite propanedioyl-CoA. First, propanedioyl-CoA is converted to propanedioyl-CoA methyl ester by a polypeptide having the activity of a S~adenosyl-L- methionine (SAM)-dep ndent methyltransferase classified, for example, under EC 2.1.1 ,197 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52), Second, propanedioyl-CoA methyl ester is enzymatically converted to a Cs _n +s aliphatic backbone via n cycles of methyl-ester shielded carbon chain elongation, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight. See FIG. 2. Each cycle m of the n cycles of methyl-ester shielded carbon chain elongation Includes: the conversion of H ₃ COC(-0)(CH ₂ ) ₂ m-5C(-0)S-CoA with acetyl-CoA to a 3-oxo-carboxyl-CoA methyl ester H ₃ COC(=0)(CH ₂ ) _2m -]C(=0)CH ₂ C(=0)S-CoA by a polypeptide having the activity of a β- ketothiolase classified, for example, under EC 2,3, 1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17) or by conversion with propanedioyl-CoA by a polypeptide having the activity of a β-k ioacyl-lacp] synthase classified, for example, under EC 2.3.1.180 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to a 3-hydroxy-carboxyl-CoA methyl ester H ₃ t¾C( ^{: )} )((¾)2 - j CH(OH)CH ₂ C(~0)S-CoA by a polypeptide having the activity of a 3-oxoacyl~[acp] reductase classified, for example, under EC L I .1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3-hydroxyacyl~CoA dehydrogenase classified, for example, under EC 1.1.1.157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 4), or a polypeptide having the activity of an acetoaceiyl-CoA reductase classified, for example, under EC 1.1 .1.36 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 3); followed by conversion to ¾COC(-0)(CH ₂ > _2m -iCH-CHC(=0)S-CoA by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1.1 19 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to H ₃ COC(=0)(CH ₂ ) ₂ m÷tC(-0)S~CoA by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyl-facp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6) or a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.38 or EC i .3.1.8 (e.g., a polypeptide having at least 50%, at least 60%>, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7). See FIG. 2.

Pathways Using NADH-Speeific Enzymes to Produce a Carboxyl-CoA Methyl Ester as a Central Precursor Leading to Difimciicma! Products

in some embodiments, a C2n+3 aliphatic backbone H ₃ COC(=0)(CH2)2n ₊ iC(= ^: 0)S-CoA ₎ also referred to as a carboxyl-CoA methyl ester, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central metabolite propanedioyl-CoA. First, propanedioyl-CoA is converted to propanedioyi-CoA methyl ester by a polypeptide having the activity of a S-adenosyl-L~ methionine (SAM)-dependent methy!transferase classified, for example, under EC 2. LI .197 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52). Second, propanedioyl-CoA methyl ester is enzymatical!y converted to a carboxyl-CoA methyl ester H ₃ CQC(~0)(C¾)2n ₊ iC(=0)S-CoA via « cycles of methyl-ester shielded carbon chain elongation, wherein « is an integer greater than or equal to one, such as, for example, one, two, three, four, live, six, seven, or eight. See FIG. 3. Each cycle m of the n cycles of methyl-ester shielded carbon chain elongation includes: the conversion of H ₃ COC{=0)(CH ₂ )2 _m .iC(-0)S-CoA with acetyl-CoA to a 3-oxo-carboxy!-CoA methyl ester l-i ₃ COC(=0)(CH ₂ )2m-iC(= ^: 0)CH2C(=0)S-CoA. by a polypeptide having the activity of a β- keiothiotase classified, for example, under EC 2.3, 1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17) or by conversion with propanedioyl-CoA by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2,3.1.180 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence any one of SEQ) ID NOs: 14-16); followed by conversion to a 3-hydroxy-carboxyl-C A methyl ester H3COC(=0)(CH2)2m- tCH(QH)C¾C( ^:::: 0)S-CoA by a polypeptide having the activity of a 3~hydroxyaeyl~CoA dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1 .1.35) (e.g., a , _ _ polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 2); followed by conversion to a 2,3- dehydrocarboxyl-CoA methyl ester H ₃ COC(==0)(CH2)2 _m -iCH=CHC(=0)S-CoA by a polypeptide having the activity of an enoyl-CoA hydratme classified, for example, under EC 4.2,1.17 (e.g., a polypeptide having at least 50%, at least 60%», at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to H ₃ COC(=0)(CH2)2ra ₊ sC( ^:::: 0)S-CoA by a polypeptide having the activity of a trans~2~enoyl- CoA reductase classified, for example, under EC 1.3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 1 15) or a polypeptide having the activity of a erioyI-[acp] reductase classified, for example, under EC 1.3, 1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46). See FIG. 3.

Pathways Using Carboxyl-CoA Methyl Esters and Carboxyl-ACP Methyl Esters as Cmtr&l Precursors to Dicarboxylic Acids

In some embodiments, a dicarbox lic acid wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor,

by conversion of a earboxyl-ACP methyl ester H ₃ €OC(=QXCH ₂ ) _? . _n+ iC( ^:::: 0)S-ACP to a monomerhyi carboxylate by a polypeptide having the activity of a ihioeslerase classified, for example, under EC 3.1.1.2 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ I D NO: 82), EC 3.1.1.5 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at. least 85% sequence homology to the amino acid sequence of SEQ ID NO: 84), or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2.21, or EC 3.1.2.27 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 93), or a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 .13 or SEQ ID NOs: 182495; followed by conversion to a dicarboxylic acid H0 ₂ C(CH ₂ ) _2n +i 0 ₂ H by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 , L I (e.g., a polypeptide having at. least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 4.

In some embodiments, a dicarboxylic acid H0 ₂ C(CH ₂ ) ₂ n+ ₁ CO ₂ H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, live, six. seven, or eight, is synthesized from the central precursor, H ₃ COC( ^:::: 0)(CH ₂ ) _{2 +!} C(=0)S-CoA, by conversion of a carboxyl-CoA. methyl ester to a monomethyl carboxylate by a polypeptide having the activity of a thioesierase classified, for example, under EC 3.1.1.2, EC 3.1.1.5, or EC 3.1 ,2.-, such as EC 3.1.2.14, EC 3.1.2.21 , or EC 3.1.2.27, or a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of any one of SE.Q ID NOs: 58-113 or SEQ ID NOs: 1 82-195; followed by conversion to a dicarboxylic acid Ηθ2€(Ό¾)2η ₊ ι C(¼H by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 50 or SEQ ID NO: 51). See FIG. 4.

in some embodiments, a dicarboxylic acid wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, H ₃ COC("0)(CH ₂ ) ₂ n ₊ iC(=0)S-CoA, by conversion of a carboxyl-CoA methyl ester i-l3COC(=0)(Ci-i ₂ )2 _! ^iC(= ^: 0)S-CoA to a monomethyl carboxylate H ₃ COC(=0)(CH ₂ )2n ₊ iC02H by a polypeptide having the activity of a CoA-transferase such as a glutaconate CoA-iransferase. classified, for example, under EC 2,8.3.12 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-!igase classified, for example, under EC 6.2.1.5; followed by conversion to a dicarboxylic acid H0 ₂ C(C3¾) ₂ n ₊ i CO ₂ H by a polypeptide having the activity of esterase classified, for example, under EC 3.1 , 1 .1 (e.g., a polypeptide having at least 50%, at least 60%, a least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See F G, 4, In some embodiments, a diearboxyllc acid wherein n is an integer greater than or equal to one, such as, for example, one, two. three, four, five, six, seven, or eight, is synthesized from the central precursor,

by conversion of a carboxyl-CoA methyl ester H ₃ COC(=0)(CH ₂ ) ₂ n ₊ iC(=0)S-CoA to a monomethyl carboxylate semialdehyde by a polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, under EC 1.2.1.1.0 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1 ,2.1 ,76, or a polypeptide having the activity of an oxogiutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion to a monomethyl carboxylate

by a polypeptide having the activity of a non-acylating NAD-dependeni aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1). a 7-oxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 24); followed by conversion to a diearboxyllc acid HOaCCCI-yan-nCOaH by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ D NO: 50 or SEQ ID NO: 51). See FIG. 4.

In some embodiments, a diearboxyllc acid H0 ₂ C(C!¾) ₂ n _* iC0 ₂ H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, by conversion of a nionomethyl carboxylate semialdehyde H3COC(=0)(CJ¼)2n+iCH(=Q) to a nionomethyl carboxylate H ₃ COC( ))(CH2 iC02H by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7-oxoheptcmoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2,1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid, sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to a dicarboxylie acid H02C(CH2)2 _f ci-iC02H by a polypeptide having the activity of an esterase classified, for example, under EC 3, 1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 4.

Pathways Using a Carboxy!-CoA Methyl Ester or a Moaomet yi Carboxylate Semialdehyde as a Centra! Precursor to an Ammocarfeoxylate

In some embodiments, an aminoearboxylate ¾M(C¾)2n ₊₂ C02H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six. seven, or eight, is synthesized from the central precursor,

by conversion of a carboxyl-CoA methyl ester ¾COC(=0)(CH ₂ )2n ₊ sC( ^:::: 0)S~CQA to a nionomethyl carboxylate semialdehyde H ₃ CQC(=0)(CH2)2a*iCH(=0) by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ I ^' D NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxogiuiarate dehydrogenase classified, for example, under EC 1 .2.1.52; followed by conversion of the monomethyl carboxylate semialdehyde H ₃ COC{=0)(CH ₂ ) _2n ÷iCH(===l-)) to a monomethyl aminocarboxylate ¾N(CH _{2 2} C( ))OCH ₃ by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.", such as EC 2.6.1 , 1 L EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1 .64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 : followed by conversion of the monomethyl aminocarboxylate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 5. n some embodiments, an aminocarboxylate H ₂ N(CH ₂ _n ₊₂ C0 ₂ H, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, ¾COC( 3)(CH ₂ ) _2n +sCH(=0), by conversion of the monomethyl carboxylate semialdehyde H ₃ COC(=0)(CH ₂ ) ₂ n+i CH(=0) to a monomethyl aminocarboxylate H ₂ N(CH ) ₂ n ₊₂ C(=0)OCH ₃ by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1 .-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1 .29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of the monomethyl aminocarboxylate H ₂ N(CH ₂ ) ₂ n ₊₂ C(=Q)OCH ₃ to ail aminocarboxylate H ₂ N(CH ₂ ) ₂ n ₊₂ C0 ₂ H by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 5. in some embodiments, an aminocarboxylate H ₂ N(CH ₂ ) _2n+2 C0 ₂ H ₅ wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, by conversion of the monomethyl carboxylate to a monomethyl carboxylate semiaidehyde by a polypeptide having the activity of a carboxylase reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at ast 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantethein transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of the monomethyl carboxylate semiaidehyde to a monomethyl aminocarboxy!ate

by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2,6.1.11. EC 2,6.1.13, EC 2.6.1.18, EC 2.6.1 .1 , EC 2.6.1.29, EC 2,6, 1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5,4.3.8 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of the monomethyl aminocarboxylate H2N(CH ₂ ):¾+2C(=0)OCH ₃ to an aminocarboxylate ¾N(CH2)2itf- ₂ C02H by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 .1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG, 5.

In some embodiments, an aminocarboxylate HaN^ loja.n ₊ aCC^H, wherein n is an integer greater tha or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, H0 ₂ C(CH2)2n+iCQ ₂ H, by conversion of the dicarboxyiic acid H02C(CH ₂ )2n+iC02H to a carboxylate semiaidehyde HOC(=0)(CH ₂ ^' _f CH(=0) by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8,7 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of the earboxyiate semialdebyde to an aminocarboxy!ate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1 .1 1 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1,64, or EC 5.4,3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181. See FIG. 5,

Path ays Using Amis¾ earboxylate, Hydroxy carboxylase, or Carboxykte Semialdebyde as Central Precursors to Diamine

In some embodiments, a diamine wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, ¾N(CH _{2 2} C0 ₂ H, by conversion of the aminocarboxylate H ₂ N(Cl¾)2n+ ₂ C02H to an aminoaldehyde HC(=0)(CH2)2n)-2 H2 by a polypeptide having the activity of a earboxyiate reductase classified, for example, under EC 1 ,2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phospkopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of the aminoaldehyde Η0( ^::: 0) Ο¾}2 _ΓΗ2 ¾ to a diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1 ,-, such as EC 2.6.1.1 , 1 1 , EC 2.6,1.13, EC 2.6.1.1 8, EC 2,6.1.19, EC 2,6, 1.29, EC 2.6,1.48, or EC 2,6.1 ,82, EC 4.1.1 ,64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181. See FIG, 6A.

The earboxyiate reductase encoded by the gene product of car and the phospkopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal di functional C4 and C5 carboxylic acids (Venkitasubramanian et ah, Enzyme and Microbial Technology, 2008, 42, 130 - 137).

In some embodiments, a diamine H2N(C¾)?. _n ÷3NH2, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, HO(C]¾) ₂ n ₊₂ 0 ₂ H (which can he produced as described in FIG. 7), by conversion of the hydroxycarboxylate HO(CH?)?.n+2C02H to an hydroxyaidehyde by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54- 57); followed by conversion of the hydroxyaidehyde HO(CH ₂ ) _2K+2 CH(~0) to a hydroxylamine HO(C¾)2n+3NH2 by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2,6.1.19, EC ^' 2.6.1.29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 : followed by conversion to an aminoaldehyde HC(=0)(CH ₂ ) ₂ n+2 I¾ by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 .1.1.- (e.g., EC 1.1.1.1, EC 1.1 .1.2, EC 1.1.1.21 , EC 1 .1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to a diamine ¾Ν(0¾)2η+3ΝΗ2 by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1.-, such as EC 2.6.1.1 1 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181. See FIG. 6 A. In some embodiments, a diamine H2N(C¾)2 _fl +3NH2, wherein » is an integer greater than or equal to one, such as, for example, one. two, three, four, five, six, seven, or eight, is synthesized from the central precursor, by conversion of an aminocarboxylate H ₂ N(CH2.)2n ₊₂ C0 ₂ H to an acetamidocarboxylate H ₃ CC(=0)NH(CH2)2n ₊ 2C02H by a polypeptide having the activity of an N- cetyltransferase such as a lysine N-acetyltransferase classified, for example, under EC 2.3.1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequenc homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to an acetamidoaldehyde H ₃ CC(=0)NH(CH2) _2ft +2CH(= )) by a polypeptide having the activity of a carboxyiate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or a least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to an aceiamidoamine H3CC(=0)NH(C¾)2n+3N¾ by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to a diamine Η ₂ Ν(0¾)2η ₊ 3ΝΗ2 by a polypeptide having the activity of an acetylputrescine deacylase classified, for example, under EC 3.5.1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45), See FIG. 6B.

In some embodiments, a diamine H ₂ N(CH ₂ ) _2S! +3NH ₂ , wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, HOC(=0)(CH2)2n ₊ iCH(=0), by conversion of the carboxyiate semi aldehyde HOC(K ⁾ )(CH ₂ ) ₂ „ ₊ iCH( )) to a dial HC(-0)(CH ₂ ) _2n+ iCH(<)) by a polypeptide having the activity of a carboxyiate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypepiide having the activity of a phosphopanietheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%» sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to an aminoaidehyde HC(=Q)(CH ₂ ):> _n +2NH2 by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1..-, such as EC 2.6.1.1 1, EC 2.6.1.1 3, EC 2.6.1.18, EC 2.6.1.19, EC 2.6, 1 .29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4.1 .1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at. least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to a diamine ¾N(CH2)2i 3 H2 by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1 .13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1 .48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypepiide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-1 81. See FIG. 6B.

Pathways Using a Ca boxylase Sesmaldehyde or a Carboxy!-CoA Methyl Ester as a Central Precursor to a Hydroxyearboxyiate

In some embodiments, a hydroxycarboxylate HO(CH2)2n _÷ 2C0 ₂ H, wherein n is an integer greater than or equal to one, such as, for example, one, two. three, four, five, six, seven, or eight is synthesized from the central precursor, H0 ₂ C(CI¾) ₂ ii ₊ iCO>H, by conversio of the dicarboxylic acid H0 ₂ C(CI¾)2n+ 1CO2H to a earboxylate semialdehyde HOC(=0)(CH2)2n+i CH^O) by a polypepiide having the activity of a earboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at. least 60%, at least 70%, or at least. 85%) sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanietheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at. least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to a hydroxycarboxylate HQ(CH2) _2n+2 CQ ₂ H by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1 ,1.- such as a 6- hydroxyhexanoate dehydrogenase classified, for example, under EC 1 .1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4~hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%>, at least 70%o, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23). See FIG. 7.

In some embodiments, a hydroxycarboxylate Η0(Ο¾) _2ΕΗ - ₂ 00 ₂ Η, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, H ₃ COC(=0)(CH2)2n+iC(=0)S-CoA, by conversion of the carboxyl-CoA methyl ester H3COC( ^:::: 0)(CH2)2; ₁ MC(= ^: 0)S-COA to a monomethyl carboxyiate semialdehyde H ₃ COC( ^:::: 0)(CH2)2 _TH -iCH( ^: =0) by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1 ,2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 18). a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of the monomethyl carboxyiate semialdehyde by a polypeptide having the activity of an esterase classified, for example, under EC 3.1, 1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to a hydroxycarboxylate Η0(Ο¾) ₂ η ₊₂ ϋ0 ₂ Η by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1. L- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypenlanoate dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 21), or a polypeptide having the activity of a 4-hydroxybuiyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23). See FIG. 7.

Pathways Using a Hydroxycarboxy!ate as a Cen ral Precursor to a Diol

in some embodiments, a diol HQCCHkhn ₊ aOH, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, HOiC^a a C^H, by conversion of the hydroxycarboxylate HO(CH ₂ ) _¾ ÷ ₂ CQ ₂ H to a hydroxyaldehyde HO(CH ₂ ) _2fl+2 CH(=0) by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID ^' NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of the hydroxyaldehyde HO(Cl¾ ₂ CH(=0) to a diol ΗΟ(ΟΗ ₂ ) _2(ν; . ₃ θΗ by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21 , EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). See FIG. 8.

Pathways using ara Amiaoearfooxylaie as a Central Precursor to a Hydroxyainine

In some embodiments, a hydroxyamine Η0(Ο¼)2η+3Ν]¾, wherein n is an integer greater than or equal to one, such as, for example, one, two, three, four, five, six, seven, or eight, is synthesized from the central precursor, H ₂ N(C¾)2rH- ₂ C0 ₂ H, by conversion of the aminocarboxyiate Η ₂ Ν(€-¾) _2Ι Η·200 ₂ Η. to an aminoaldehyde by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2,7.8,7 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of the aminoaldehyde HC( ^:::: 0)(CH2)2n ₊ 2 H2 to a hydroxyamme HQ(CH2)2n÷3 ¾ by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1 , 1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1 .1.21, EC 1.1.1.61 , or EC 1.1 .1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). See FIG, 6A.

C$ Bioehemkal Pathways

Pathways Using NADPH-Spedfk Enzymes to Produce Penianedioyl-jacp] Methyl Ester as a Centra! Precursor Leading to C _¾ Building Blocks

In some embodiments, pentanedioyl-[acp] methyl ester is synthesized from the central metabolite propanedioyl~[acp! via one cycle of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-jacp] to propanedioyl~[acp] methyl ester by a polypeptide having the activity of a S-adenosyl-L-methionine (SAM) -dependent meihyltramj erase classified, for example, under EC 2.1.1.197 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52); followed by conversion with propanedioyl-[acp] to 3~oxo-pentanedioyl-[aep] methyl ester by a polypeptide having the activity of a f]~ketoacyl~[acp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1 ,41 , EC 2.3.1.179, or EC 2.3.1.180) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3-hydroxy-pentanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3-oxoacyl-facp] reductase classified, for example, under EC 1.1.1.100 (e.g., a polypeptide having at least 50%, at least. 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3- dehydropentanedioyl-[acpj methyl ester by a polypeptide having the activity of a 3~ h droxyacyl~[ acp ] dehydratase classified, for example, under EC 4,2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to pentanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl-facp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at. least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6), See FIG. 33.

Pathways Using NADPH-Specific Enzymes to Produce Peetasiedioyl-CoA Methyl Ester m a Central Precursor Leadin to C§ Building Blocks

in some embodiments, pentanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via one cycle of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to propanedioyl-CoA methyl ester by a polypeptide having the activity of a S-adenosyl~L-methionine (SAM)-dependent methyltr nsferase classified, for example, under EC 2, 1.1.197 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of SEQ ID NO; 52); followed by conversion with acetyl-CoA to 3-oxo-pentanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketothiolase classified, for example, under EC 2.3.1 ,16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17) or by conversion with propanedioyl-CoA by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.180 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3-hydroxy-pentanedioyl-CoA methyl ester by a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified, for example, under EC 1 ,1 , 1.100 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1 , L I .157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4). or a polypeptide having the activity of an acetoaceiyl-CoA reductase classified, for example, under EC 1 , 1.1.36 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydropentanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyUCoA hydraiase classified, for example, under EC 4,2.1.1 19 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to pentanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3. L- such as an enoyl-[acp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6) or a irans-2-enoyl-CoA reductase classified, for example, under EC 1 .3.1.38 or EC 1 ,3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7), See FIG. 34.

Pathways Using ADH-Spedfk Enzymes to Produce Pe»tanedioyi~CeA Methyl Ester as a Central Preem's r Leading to€5 Building Blocks

In some embodiments, pentanedioyi-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via one cycle of methyl -ester shielded carbon chain elongation by conversion of propanedioyl-CoA to propanedioyl-CoA methyl ester by a polypeptide having the activity of a S-adenosyl-L-methionine (SA.M)~dependent methyliransferase classified, for example, under EC 2.1.1.1 97 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 52); followed by conversion with acety -CoA to 3-oxo-pentanedioyl~CoA methyl ester by a polypeptide having the activity of a β-ketothiolase classified, for example, under EC! 2.3,1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 17) or by conversion with propanedioyi-CoA by a polypeptide having the activity of a β-ketoacyl-facpj synthase classified, for example, under EC 2.3.1.180 (e.g., a polypeptide having at least 50%, at least 60%. at least. 70%, or at least 85% sequence homology to the amino acid sequence any one of SEQ ID NOs: 14-16); followed by conversion to 3~hydroxy~pentanedioyl~CoA methyl ester by a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g., EC LI .1.35) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3 -dehydropentanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1.17 (e.g., e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to pentanedioyi-CoA methyl ester by a polypeptide having the activity of a trans- 2-enoyl-CoA reductase classified, for example, under EC 1 ,3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 114 or SEQ ID NO: 115) or a polypeptide having the activity of a enoyl-facp] reductase classified, for example, under EC 1.3.1 ,9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85¾> sequence homology to the amino acid sequence of SEQ ID NO: 46). See FIG. 35.

Pathways Using Penianedloyl-CoA Methyl Ester or Pentanedioy!-facp] Methyl Ester as Central Precursors to Pentanedioic Acid

In some embodiments, pentanedioic acid is synthesized from the central precursor, pentanedioyl-[acp] methyl ester, by conversion of pentanedioyl-[aep] methyl ester to monomethyl pentanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3,1.1 ,2, EC 3.1 , 1.5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2.21, or EC 3.1 .2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to pentanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at. least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

in some embodiments, penianedioic acid is synthesized from the centra! precursor, pentanedioyl-CoA methyl ester, by conversion of perstanedioyl-CoA methyl ester to monomethyl pe tanedioate by a polypeptide having the activity of a Ihioesterase classified, for example, under EC 3.1.1.2, EC 3.1.1 .5, or EC 3.1.2.-, such as EC 3, 1.2.14, EC 3.1.2.21 , or EC 3 J .2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to penianedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, penianedioic acid is synthesized from the central precursor, pentanedioyl-CoA. methyl ester, by conversion of pentanedioyl-CoA methyl ester to monomethyl pentanedioate by a polypeptide having the activity of a CoA-transferase such as a glutaconate CoA-transferase classified, for example, under EC 2.8.3.12 (e.g., a polypeptide having at least 50%, at least. 60%, at. least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-iigase classified, for example, under EC 6.2,1.5; followed by conversion to penianedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1 ,1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, penianedioic acid is synthesized from the central precursor, pentanedioyl-CoA methyl esier, by conversion of pentanedioyi-Co A methyl ester to methyl 5~ oxopentanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, under EC 1 ,2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 8), a polypeptide having at least 50%, at least 60%, at least 70'%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion to monomethyl pentanedioate by a polypeptide having the activity of a a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7-oxoheptanoaie dehydrogenase classified, for example, under EC 1.2.1 ,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO; 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2,1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to pentanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, pentanedioic acid is synthesized from the central precursor, methyl 5-oxopentanoate, by conversion of methyl 5-oxopentanoate to monomethyl pentanedioate by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; Π), a 7- oxoheptanoate dehydrogenase classified, for example, under EC I.2.I .- (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%>, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1 ,2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to pentanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3, 1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 ).

Pathways Using Pentanedioyi-CoA Methyl Ester or Methyl S-Oxopeistanoate as a Central Precursor to 5~Aminop«staEoate

in some embodiments, 5-aminopentanoate is synthesized from the central precursor, pentanedioyl-CoA methyl ester, by conversion of pentanedioyi-CoA methyl ester to methyl 5- oxopentanoate by a polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ D NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 5-oxopentanoate to monomeihyl 5- aminopentanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2,6.1.11 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6, 1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16438 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 5-aminopentanoate to 5~aniinopentanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

in some embodiments, 5-aminopentanoate is synthesized from the central precursor, methyl 5-oxopentanoate, by conversion of methyl 5-oxopentanoate to monomethyl 5- aminopentanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6,1 .29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1 ,64, EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16438 or SEQ ID NOs: 167481 ; followed by conversion of monomet yl 5-aminopentanoate to 5-aminopentanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%>, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 5-aminopentanoate is synthesized from the central precursor, monomethyl pentanedioate, by conversion of monomethyl pentanedioate to methyl 5- oxopentanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed byconversion of methyl 5-oxopentanoate to monomethyl 5-aminopentanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6,1.1 1 , EC 2,6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1 ,1.64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 5- aminopentanoate to 5-aminopentanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 5-aminopentanoate is synthesized from the central precursor, penianedioic acid, by conversion of penianedioic acid to 5-oxopentanoate by a polypeptide fiaving the activity of a carboxylate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanietheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having a least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 5~ oxopentanoate to 5-aminopentanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-1 81.

Pathways Usmg S-Amraopesstanoate, S-Hydroxypenianoate, or S-Oxopentanoafe as Central Precursors to Peiitarie~l,5-Di¾miiie

in some embodiments, pentane-l,5-diamine is synthesized from the central precursor, 5-aminopentanoate, by conversion of 5-aminopentanoate to 5-aminopentanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanietheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 5- aminopentanal to pentane-l,5-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1 such as EC 2.6,1.11, EC 2.6.1 , 13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1 .29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4, 1 .1.64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

The carboxylate reductase encoded by the gene product of car and the phosphopanietheine transferase enhancer npt or sfp has broad substrate specificity, including terminal Afunctional C4 and C5 carboxylic acids (Venkitasubramanian et al, Enzyme and Microbial Technology, 2008, 42, 130 - 137).

In some embodiments, pentane- 1 ,5-diamine is synthesized from the central precursor, 5-hydroxypentanoate, by conversion of 5-hydroxypentanoate to 5-hydroxypentanal by a polypeptide having the activity of a carboxyiate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SliQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid, sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 5~ hydroxypentanal to 5-ammopentanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2,6.1.13, EC 2.6.1 ,1 8, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 : followed by conversion to 5-aminopentanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1 ,1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC i .1 ,1.21, EC 1 ,1.1.61, or EC 1.1.1 , 184) (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to pentane- 1 ,5-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1 , EC 2.6.1.13, EC 2.6.1 .18, EC 2.6.1 .19, EC 2,6.1.29, EC 2,6.1.48, or EC 2.6, 1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%:·, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167481.

In some embodiments, pentane- 1,5 -diamine is synthesized from the central precursor, 5-arainopentanoate, by conversion of 5-aminopentanoate to N5-acetyl-5-aminopentanoate by a polypeptide having the activity of an N-acetyltransferase such as a lysine N- aceiyltransferase classified, for example, under EC 2,3.1.32 (e.g., a polypeptide having at least 50%, at leasi 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 53); followed by conversion to N5-acetyl-5-aminopentanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8,-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to N5~ acetyl- 1 ,5-diaminopentane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2,6.1.11 , EC 2.6,1.13, EC 2,6.1.18, EC 2.6, 1 .19, EC 2.6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to pentane- 1 ,5-diamine by a polypeptide having the activity of an acetylpulrescine deacylase classified, for example, under EC 3.5.1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, pentane- 1 ,5-diamine is synthesized from the central precursor, 5-oxopentanoate, by conversion of 5-oxopentanoate to 1 ,5-pentanedial by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at. least 50%, at least 60%, at least 70%», or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 5- aminopentanal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 : followed by conversion to peniatie- ί ,5-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6, 1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SE.Q ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

Pathways Using Pentanedioic Acid or Pentanedioyi-CoA Methyl Ester as a Central Precursor to 5-HydroxypeEtanoate

In some embodiments, 5-hydroxypentanoate is synthesized from the central precursor, pentanedioic acid, by conversion of pentanedioic acid to 5-oxopentanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetkeine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 5- hydroxypenianoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6~hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxyp ntanoate dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a polypeptide having the activity of a 4~hydroxybutyraie dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of SEQ ID NO: 23), in some embodiments, 5-hydroxypentanoate is synthesized from the central precursor, pentanedioyl~CoA methyl ester, by conversion of pentanedioyl-CoA methyl ester to methyl 5- oxopentanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2, 1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 , a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1,2.1,76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2,1.52; followed by conversion of methyl 5-oxopentanoate to 5 -oxopentanoaie by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 5-hydroxypentanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1 , 1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase classified, for example, under EC 1 ,1 , l .~ (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxybulyrate dehydrogenase (e.g., a polypeptide having at least 5G%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23).

Pathways Using S-Hydr xypentsnoate as a Central Precursor to l,5~Penianediol

in some embodiments, 1.5-pentanediol is synthesized from the central precursor, 5- hydroxypentanoate, by conversion of 5-hydroxypentanoate to 5-hydroxypentanai by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ΪΙ) NOs; 54-57); followed by conversion of 5- hydroxypentanal to 1,5-pentanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 .1 , 1.- (e.g., EC 1.1.1.1, EC 1, 1 .1.2, EC L I .1.21. EC 1.1.1 .61 , or EC 1.1 .1.1 4) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). Pathways Using S-Aminopeatanoate as a Central Precursor to S-Amiaopeiiian l

In some embodiments, 5-aminopentanol is synthesized from the central precursor, 5~ aminopentanoate, by conversion of 5-arainopentanoate to 5-aminopentanal by a polypeptide having the activity of a carhoxylate reductase classiiied, for example, under EC ί .2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%;, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion of 5- aminopentanal to 5-aminopentanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 , 1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21 , EC 1.1 .1.61 , or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). C ₇ Biochemical Pathways

Pathways using NADPH-Specific Enzymes to Produce Heptanedioyl-[acp] Methyl Ester as a Central Precursor Leading to C _? Building Blocks

In some embodiments. heptanedioyl~[acp] methyl ester is synthesized from the central metabolite propanedioyl~[acp] via two cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-[acp] to pentanedioyi-f acp] methyl ester as described above; followed by conversion to 3-oxo-heptanedioyl-[acp] methyl ester by a polypeptide having the activity of a β-ketoacyi-facp] synthase classified, for example, under EC 2.3. L- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3~hydroxy~heptanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3~oxoacyi-[acp] reductase classified, for example, under EC 1.1.1 , 100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3-dehydroheptanedioyl-[aep] methyl ester by a polypeptide having the activity of a 3-hydroxyacyl-[acp] dehydratase classified, for example, under EC 4,2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to heptanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl-facp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ D NO: 6), See FIG. 33.

Pathways Using NADPB-Spedfie Eraaymes to Produce Heptanedioyi-CoA Methyl Ester as a Central Precursor Leading to C _? Building Blocks

in some embodiments, heptanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via two cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to pentanedioyl-CoA methyl ester as described above; followed by conversion to 3~oxo-heptanedioyl~CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2,3.1.16 (e.g., a polypeptide having at least 50%, at least 60%), at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3-hydroxy-heptanedioyl- CoA methyl ester by a polypeptide having the activity of a 3-oxo cyl~[acp] reductase classified, for example, under EC 1 , 1.1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3-hydrox acyl-CoA dehydrogenase classified, for example, under EC 1 , 1.1.157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4), or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC 1 .1.1 ,36 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydroheptanedioyhCoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1.1 19 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to heptanedioy] -CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyi-f cp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 6) or a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.38 or EC 1.3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7). See FIG. 34,

Pathways Using NADH-Specific Enzymes to Produce Heptanedioy!-CoA Methyl Ester as a Centra! Precursor Leading to C ₇ Banding Blocks

In some embodiments, heptanedioy I -CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via two cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to pentanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo-heptanedioyhCoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1 ,41 or EC 2.3.1 , 179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothioiase classified, for example, under EC 2.3,1 , 16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3~hydroxy-heptanedioyl- CoA methyl ester by a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1.1 ,135 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3-dehydroheptanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1 ,17 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to heptanedioyl-CoA methyl ester by a polypeptide having the activity of a trans- 2-enoyl-CoA reductase classified, for example, under EC 1.3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 1 15) or a polypeptide having th activity of a enoyi-facp] reductase classified, for example, under EC 1.3.1 ,9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%*, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46). See FIG. 35.

Pathways Using Heptanedioyl-CoA Methyl Ester or Heptaiiedioyl-[acpj Methyl Ester as Central Precursors to Heptsmedioie Add

In some embodiments, heptanedioic acid is synthesized from the central precursor, hcptanedioyI-[acp] methyl ester, by conversion of heptanedioyl-[acp] methyl ester to monomethyl heptanedioate by a polypeptide having the activity of a tkioesterase classified, for example, under EC 3.1.1.2, EC 3,1.1.5, or EC 3.1.2.-, such as EC 3, 1.2.14, EC 3,1 .2.21 , or EC 3,1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to heptanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG, 36. In some embodiments, heptanedioic acid is synthesized from the central precursor, heptanedioyl-CoA methyl ester, by conversion of heptanedioyl-CoA methyl ester to monomethyl heptanedioate by a polypepiide having the activity of a thioesterase classified, for example, under EC 3.1.1.2, EC 3.1 ,1.5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2,21, or EC 3.1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to heptanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 36.

In some embodiments, heptanedioic acid is synthesized from the central precursor, heptanedioy!-CoA methyl ester, by conversion of heptanedioyl-CoA methyl ester to monomethyl heptanedioate by a polypeptide having the activity of a CoA-irartsfer se such as a glutaconate CoA-transferase classified, for example, under EC 2,8.3.12 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-iigase classified, for example, under EC 6.2.1.5; followed by conversion to heptanedioic acicl by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 36.

In some embodiments, heptanedioic acid is synthesized from the central precursor, heptanedioyl-CoA methyl ester, by conversion of heptanedioyi-CoA methyl ester to methyl 7~ oxoheptanoaie by a polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, under EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypepiide having at least 50%, at least 60%, at least 70%o, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semia!dehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1 ,52; followed by conversion to monomethyl heptanedioate by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 11), a 7~oxoheptanoate dehydrogenase classified, for example, under EC 1 ,2.1.- (e.g., a polypeptide having at least 50%, at least 60%», at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoaie dehydrogenase classified, for example, under EC 1.2,1 ,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to heptanedioic acid by a polypeptide having the activity of an esierase classified, for example, under EC 3.1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 36.

In some embodiments, heptanedioic acid is synthesized from the central precursor, methyl 7-oxohepianoate. by conversion of methyl 7-oxoheptanoate to monomethyl heptanedioate by a polypeptide having the activity of a non-acylating NAD-dependeni aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: I I), a 7- oxohepianoate dehydrogenase classified, for example, under EC 1 ,2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to heptanedioic acid by a polypeptide having the activity of an esierase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, ai least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 36.

Pathways Usmg Beptanedioyl-CoA Methyl Ester or Methyl T-OxobeptaHso&te as a Central Precursor to 7-Aminoheptaaoate

In some embodiments, 7-aminoheptanoate is synthesized from the central precursor, heptanedioyl-CoA methyl ester, by conversion of heptanedioyl-CoA methyl ester to methyl 7~ oxoheptanoaie by a polypeptide having the activity of an acet lating aldehyde dehydrogenase classified, for example, EC 1.2.1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxogiutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 7-oxoheptanoate to monomethyl 7- aminoheptanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6,1.13, EC 2.6.1,18, EC 2.6.1 ,19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6.1 ,82, EC 4.1 ,1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ II) NOs: 116-138 or SEQ ID NOs: 167-181 : followed by conversion of monomethyl 7-aminoheptanoate to 7-aminoheptanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence. homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 37.

In some embodiments, 7-aminoheptanoate is synthesized from the central precursor, methyl 7-oxoheptanoate, by conversion of methyl 7~oxohep†.anoate to monomethyl 7- aminoheptanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6.1 , 1 1, EC 2.6.1.13, EC 2,6.1.18, EC 2,6.1.19, EC 2,6.1.29, EC 2,6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5.4.38, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 7-aminoheptanoate to 7-aminoheptanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 .1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 37.

In some embodiments, 7-aminoheptanoate is synthesized from the central precursor, monomethyl heptanedioate, by conversion of monomethyl heptanedioate to methyl 7- oxoheptanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of methyl 7-oxoheptanoate to monomethyl 7-aminoheptanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2,6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-338 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 7- aminoheptanoate to 7-aminoheptanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). See FIG. 37.

In some embodiments, 7-aminoheptanoate is synthesized from the central precursor, heptanedioic acid, by conversion of heptanedioic acid to 7-oxoheptanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 7- oxoheptanoate to 7-aminoheptanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.1.9, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181, See FIG. 37. Pathways Using 7-Aniiffioheptanossie, 7-HydroxyheptaEoate, or 7-Oxoheptanoate m Central Precursors to Heptajie-I,7-Diamme

In some embodiments, heptane- 1 ,7-diamine is synthesized from the central precursor, 7-aminoheptanoate, by conversion of 7~aminoheptanoate to 7-aminoheptanaI by a polypeptide having the activity of a carboxy!ate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a pkosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 7- aminoheptanal to heptane- 1 ,7-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2,6.1 , 11, EC 2.6,1 , 13, EC 2.6.1.18, EC 2.6, 1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4.1 ,1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs; 167-181. See FIG. 38.

The carboxylaie reductase encoded by the gene product of car and the pkosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunciional C4 and C5 carboxylic acids (Venkitasubramanian et ah, Enzyme and Microbial Technology, 2008, 42, 130 - 137).

In some embodiments, heptane- 1 ,7-diamine is synthesized from the central precursor, 7-hydroxyhepianoate (which can be produced as described in FIG. 39), by conversion of 7- hydroxyheptanoate to 7-hydroxyheptanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7,8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 7-hydroxyheptanal to 7-aminoheptanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2,6.1.1 1 , EC 2.6.1 ,13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2.6.1 .82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%o. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to 7-aminoheptanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1 ,1, EC 1 .1.1.2, EC 1.1.1 ,21, EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to heptane- 1,7-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1.-, such as EC 2.6.1.1 1 , EC 2,6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%), at least 70%, or at least 85%) sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181. See FIG. 38,

in some embodiments, heptane- 1,7-diamine is synthesized from the central precursor, 7-aminoheptanoate, by conversion of 7-aminoheptanoate to N7-acetyl-7-aminoheptanoate by a poiypeptide having the activity of an N~aceiyliransferase such as a lysine N- acetyltransferase classified, for example, under EC 2.3.1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to N7-acetyl-7-aminolieptanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at [east 70%;, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a plwsphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50% , at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to N7- acetyl- 1 ,7-diaminoheptane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6. L I E EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to heptane- 1,7-diamine by a polypeptide having the activity of an acetylpntre seine de c lase classified, for example, under EC 3.5.1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45). See FIG, 38.

In some embodiments, heptane- 1,7-diamine is synthesized from the central precursor,

7-oxoheptanoate, by conversion of 7-oxoheptanoate to 1,7-heptanedial by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7.8.-, such as 2.7,8,7 (e.g., a polypepiide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 7- aminoheptanal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11. EC 2.6.1.13, EC 2.6.1 .18, EC 2.6.1.19, EC 2.6,1 ,29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypepiide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by ^¬ conversion to heptane- 1, 7~diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181. See FIG. 38.

Pathwa s Using Heptanediok add or Heptanedioyl-CoA Methyl Ester as a Central Precursor to T-Hydroxyheptasioate

In some embodiments, 7-hydroxyheptaiioate is synthesized from the central precursor, heptanedioic acid, by conversion of heptanedioic acid to 7-oxoheptanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 7~ hydroxyhepianoate by a polypeptide having the activity of a dehydrogenase classified, for example,, under EC 1.1.1.- such as a 6~hydroxyhexanoate dehydrogenase classified, tor example, under EC 1.1 ,1 ,258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5~hydroxypentanoate dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxyhutyraie dehydrogenase (e.g., a polypepiide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 23). See FIG. 39.

In some embodiments, 7-hydroxyheptanoate is synthesized from the central precursor, heptanedioyl-CoA methyl ester, by conversion of he tanedioyl-CoA methyl ester to methyl 7- oxoheptanoate by a polypeptide having the activity of an aceiylating aldehyde dehydrogenase classified, for example, EC 1 ,2.1 .10 (e.g., a polypeptide having at least 50%, at least 60%), at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 9, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1 ,2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 7-oxoheptanoate to 7-oxoheptanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3. 1 .1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 5 1); followed by ^¬ conversion to 7~hydroxyheptanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1 , 1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1 .1 .258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%s sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a polypeptide having the activity of a 4-hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%*, at least 60%, at least 7Q%>, or at least 85%s sequence homology to the amino acid sequence of SEQ ID NO: 23). See FIG. 39,

Pathways Using 7-Hydroxyheptanoaie as a Central Precursor to I ₅ 7-Hep†s¾Eediol

In some embodiments, 1 ,7-heptanediol is synthesized from the central precursor, 7- hydroxyheptanoate, by conversion of 7-hydroxyheptanoate to 7-hydroxyheptanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopaniethem ^' e transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at [east 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 7- hydroxyheptanai to 1 ,7-heptanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1 .- (e.g., EC 1.1.1.1 , EC 1.1.1 .2, EC 1.1.1.21 , EC 1 .1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). See FIG. 40.

Pathways Using 7~Amirsoheptai¾oaie as a C ntra? Precursor to 7-Amiiiohepiasiol

in some embodiments, 7-aminoheptanol is synthesized from the central precursor, 7- aminoheptanoate, by conversion of 7-ammoheptanoate to 7-aminoheptanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 7- aminoheptanal to 7-aminoheptanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1 .1.- (e.g., EC 1.1.1.1, EC 1 ,1 , 1.2, EC 1.1.1.21 , EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%», or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). See FIG. 38. C9 Biochemical Pathways

Pathways Using NADPH-Specific Enz mes to Produce Nonaiiedioyl-[acp] Methyl Ester as a Central Precursor L ading to C Building Blocks

In some embodiments, nonanedioyl-[acp] methyl ester is synthesized from the central metabolite propanedioyi-[acp] via three cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-[aep] to heptanedioyl~[acp] methyl ester as described above; followed by conversion to 3~oxo~nonanedioyl~[acp] methyl ester by a polypeptide having the activity of a β-ketoacyl-facpj synthase classified, for example, under EC 2.3,1 .- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3-hydro y-nonanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3~oxoacyl-[acp] reductase classified, for example, under EC 1 ,1 , 1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3~dehydrononanedioy ~[acpj methyl ester by a polypeptide having the activity of a 3~hydroxyacyl-[acp] dehydratase classified, for example, under EC 4.2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to nonanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl-facp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Using NADPH-Specific Enzymes to Produce Nonanedioyi-CoA Methyl Ester as a Centra! Precursor Leading to Q Building Blocks

In some embodiments, nonanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via three cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyi-CoA to heptanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo-nonanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 23.1,41 or EC 2,3, 1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2,3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-nonanedioyl-CoA methyl ester by a polypeptide having the activity of a 3-oxoacyl- [acp] reductase classified, for example, under EC 1 ,1.1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 5). a polypeptide having the activity of a 3-hydroxyacyi-CoA dehydrogenase classified, for example, under EC 1 , 1.1.157 (e.g., a polypeptide having at least 50%, at least. 60%, at least. 70%>, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO; 4), or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC 1.1 .1.36 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydrononanedioyhCoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydrolase classified, for example, under EC 4.2.1.1 19 (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to nonanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyi-[acp] reductase classified, for example, under EC 1.3.1 .10 (e.g., a polypeptide having at least 50%o, at least 60%, at least 70%, or at least 85% sequenc homology to the amino acid sequence of SEQ ID NO: 6) or a irans~2-enoyl~CoA reductase classified, for example, under EC 1.3.1 ,38 or EC 1 .3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7).

Pathways Using NABH-Specifk Eraaymes to Produce Nonanedioyl-CoA Methyl Ester as Ά Central Precursor Leading to C9 Building Blocks

in some embodiments, nonanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via three cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to heptanedioyl-CoA methyl ester as described above; followed by conversion to 3~oxo-nonanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothiolase classified, for example, under EC 2.3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 17); followed by conversion to 3- hydroxy-nonanedioyl-CoA methyl ester by a. polypeptide having the activity of a 3- hy roxyacyl-CoA dehydrogenase classified, for example, under EC 1 , 1. 1.35 (e.g., a polypeptide having at least 50%o, at least 60%, a least 70%, or at least 85%s sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3- dehydrononanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydrolase classified, for example, under EC 4.2,1.17 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to nonanedioyl-CoA methyl ester by a polypeptide having the activity of a tr ns-2-enoyl-CoA reductase classified, for example, under EC 1 .3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 114 or SEQ ID NO: 1 1 5) or a polypeptide having the activity of a enoyi-facp] reductase classified, for example, under EC 1.3, 1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46).

Pathways Using Nonanedioyl-CoA Methyl Ester or Nonanedioyi-j cpj Methyl Ester as Central Precursors to onanedioic Acid

in some embodiments, nonanedioic acid is synthesized from the central precursor, nonanedioyl-[acp] methyl ester, by conversion of nonanedioyI-[acp] methyl ester to monomethyl nonanedtoate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1,1.2, EC 3.1.1.5, or EC 3.1.2.-, such as EC 3.1 .2.14, EC 3.1.2.21, or EC 3. 1 .2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to nonanedioie acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonanedioie acid is synthesized from the central precursor, nonanedio l-CoA methyl ester, by conversion of nonanedioy!-CoA methyl ester to monomethyl nonanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3. L i .2, EC 3.1.1.5, or EC 3.1 .2.-, such as EC 3.1.2.14, EC 3.1.2.21, or EC 3,1.2.27, or a polypeptide having at least 50%, at least 60%, at. least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to nonanedioie acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonanedioie acid is synthesized from the central precursor, nonanedioyl-CoA methyl ester, by conversion of nonanedioyl-CoA methyl ester to monomethyl nonanedioate by a polypeptide having the activity of a CoA-transferase such as a ghiiaccmaie CoA-transferase classified, for example, under EC 2.8.3, 12 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-ligase classified, for example, under EC 6,2.1.5; followed by conversion to nonanedioie acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonanedioie acid is synthesized from the central precursor, nonanedioyl-CoA methyl ester, b conversion of nonanedioyl-CoA methyl ester to methyl 9- oxononanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, under EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion to monomethyl nonanedioate by a polypeptide having the activity of a non-acylating NAD~dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 1 1), a 7-oxoheptanoate dehydrogenase classified, for example, under EC 1.2,.! ,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to nonanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 , 1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO; 1).

In some embodiments, nonanedioic acid is synthesized from the central precursor, methyl 9-oxononanoate, by conversion of methyl 9-oxononanoate to monomethyl nonanedioate by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at leas 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7~ oxoheplanoate dehydrogenase classified, for example, under EC 1 ,2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1 ,2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to nonanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%. a least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

Pathways Using onanedloyl-CoA Methyl Ester or Methyl 9~Oxononanoate as a Central Precursor to 9-Amiii nonaBoate

In some embodiments, 9-aminononanoate is synthesized from the central precursor, nonanedioyl-CoA methyl ester, by conversion of nonanedioyl-CoA methyl ester to methyl 9- oxononanoate by a polypeptide having the activit of an acetylatmg aldehyde dehydrogenase classified, for example, EC 1 ,2.1 ,10 (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 8), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1 ,76, or a. polypeptide having the activity of an oxoglittarate dehydrogenase classified, for example, under EC 1 .2.1.52; followed by conversion of methyl 9-oxononatioate to monomethyl 9- aminononanoate by a polypeptide having the activity of an aminotransferase classified, for example, under E EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6. L 13, EC 2,6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: .167-181 ; followed by conversion of monomethyl 9-aminononanoate to 9-aminononanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3, 1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 9-aminononanoate is synthesized from the central precursor, methyl 9-oxononanoate, by conversion of methyl 9-oxononanoate to monomethyl 9~ aminononanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monoraethyl 9-aminononanoate to 9-aminononanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 50 or SEQ ID NO: 51).

In some embodiments, 9-aminononanoate is synthesized from the central precursor, monomethyl nonanedioate, by conversion of monomethyl nonanedioate to methyl 9- oxononanoate by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 5G%>, at least 60%, at least 70%, or at least 85 % sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of methyl 9-oxononanoate to monomethyl 9-aminononanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.1 8, EC 2,6.1.19, EC 2.6.1.29, EC 2.6,1 .48, or EC 2.6.1.82, EC 4,1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ D NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 9~ aminononanoate to 9-aminononanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having a least 50%, at least 60%, at least. 70%, or at least 85%t sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 9-aminononanoate is synthesized from the central precursor, nonanedioic acid, by conversion of nonanedioic acid to 9-oxononanoale by a polypeptide having the activity of a carboxylaie reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs; 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 54-57); followed by conversion of 9- oxononanoate to 9-aminononanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6,1 .13, EC 2.6.1.18, EC 2.6.1 , 19, EC 2.6,1.29, EC 2.6.1.48, or EC 2.6.1 .82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 1 16-138 or SEQ ID NOs: 167-1 81.

Pathways Using 9-Amin si !ja¾oate, 9-Hydroxynersanoaie, or 9~Gxosions¾raoale as Central Precursors to NoiiiiBe-i,9-Dkmirie

In some embodiments, nonane-!,9-diamine is synthesized from the central precursor, 9-aminononanoate, by conversion of 9-aminononanoate to 9-aniinononanal by a polypeptide having the activity of a carboxylaie reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 9- aminononanal to nonane-1 ,9-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2,6.1 ,11, EC 2.6,1.13, EC 2,6.1 ,1 8, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6, 1.82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

The carhoxylate reductase encoded by the gene product of car and the phosphopanteiheine transferase enhancer npt or s/p has broad substrate specificity, including terminal difunctional€4 and C5 carboxylic acids (Venkitasubramanian et ah, Enzyme and Microbial Technology, 2008, 42, 130 ^■■■ · 137),

in some embodiments, nonane-l,9-diamine is synthesized from the central precursor, 9-hydroxynonanoate. by conversion of 9-hydroxynonanoate to 9~hydroxynonanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 9- hydroxynonanal to 9-aminononanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6, 1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1 .82, EC 4.1.1.64, or EC 5.4,3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to 9-aminononanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1 ,1 ,- (e.g., EC 1.1 .1.1, EC L I .1.2, EC 1.1.1.21 , EC 1.1.1.61 , or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to nonane-1 ,9-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6, 1 , 1 1 , EC 2,6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1 ,29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1 , 1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181. In some embodiments, nonane- 1 ,9-di.aimne is synthesized from the central precursor, 9-aminononanoate, by conversion of 9-aminononanoate to N9~aeetyl~9-arninononanoate by a polypeptide having the activity of an N-acetyltransferase such as a lysine N-acetyliransferase classified, for example, under EC 2,3, 1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to N9-acetyl-9-aminononanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to N9-acetyl-l,9- diaminononane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1.-, such as EC 2.6.1.1 1, EC 2.6, 1.13, EC 2,6.1.18, EC 2,6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4,1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to nonane- 1 ,9-diamine by a polypeptide having the activity of an acetylputrescine deacylase classified, for example, under EC 3.5.1.62 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, nonane- 1,9-di amine is synthesized from the central precursor, 9-oxononanoate, by conversion of 9-oxononanoate to 1,9-nonanedial by a polypeptide having the activity of a carhoxy!ate reductase classified, for example, under EC 1,2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8.7 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any o of SEQ ID NOs: 54-57); followed by conversion to 9- ammanonana] by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to nonane-l,9~diamme by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6,1 , 1 1 , EC 2,6.1.13, EC 2.6.1.1 8, EC 2.6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1.82, EC 4.1 ,1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

Pathways Using Nonanedioic Acid or Nonanedioyl-CoA Methyl Ester as a Central Pr cursor to 9~Hydro¾yiiona at£

In some embodiments, 9-hydroxynonanoate is synthesized from the central precursor, nonanedioic acid, by conversion of nonanedioic acid to 9-oxononanoate by a polypeptide having the activity of a carbox late reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 9- hydroxynonanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1 , 1.1.- such as a 6-hydrox hexanoate dehydrogenase classified, for example, under EC 1 ,1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentano te dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxyhutyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23).

In some embodiments _. , 9-hydroxynonanoate is synthesized from the central precursor, nonanedioyl-CoA methyl ester, by conversion of nonanedioyl-CoA methyl ester to methyl 9- oxononanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1 .2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 9-oxononanoate to 9-oxononanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 9-hydroxynonanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1 ,1 .- such as a 6~hydroxyhexanoate dehydrogenase classified, for example, under EC LI .1.258 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydmxypentanoate dehydrogenase classified, for example, under EC 1,1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydraxyhiUyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23). Pathways Using 9-llydroxys5 ?xanoate as a Central Precursor to 1,9-NonaKedioi

In some embodiments, 1,9-nonanedioi is synthesized from the central precursor, 9- hydroxynonanoate, by conversion of 9-hydroxynonanoaie to 9-hydroxynonanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ΪΪ ⁾ NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of an one of SEQ ID NOs: 54-57); followed by conversion of 9- hydroxynonanal to 1,9-nonanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1 .1.1 ,1 , EC 1.1.1.2, EC 1.1.1 .21 , EC 1 .1 .1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 20-23).

Pathways Using 9-AminoKOi¾aaoaie as a Central Precursor to 9~Ami¾onoiiaBoi

In some embodiments, 9-aminononanol. is synthesized from the central precursor, 9- aminononanoate, by conversion of 9-aminononanoate to 9~aminononanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homolog to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a. phosphopantetheine transferase classified, for example, under EC 2,7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 9- aminononanai to 9-aminononanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 , 1.1.— (e.g., EC 1.1.1.1 , EC 1.1.1.2, EC 1.1 ,1.21 , EC 1.1.1.61, or EC 1 , 1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Cn Biochemical Pathways

Pathways Using NADPH-Specific Enzymes to Produce Undecanedioyl-[acp] Methyl Ester m a Central Precursor Leading to Cn Bsiildisig Blocks

In some embodiments, undecanedioyl-[acp] methyl ester is synthesized from the central metabolite propanedioyl-[aep] via four cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-f acp] to nonanedioyI-[acp] methyl ester as described above: followed by conversion to 3 -oxo-undecanedioyl-[acpJ methyl ester by a polypeptide having the activity of a £i~keioacyi~[acpj synthase classified, for example, under EC 2,3.1.- (e.g., EC 2,3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having a least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 14-16); followed by conversion to 3-hydroxy-undecanedioyl-[acpJ methyl ester by a polypeptide having the activity of a 3-oxoacyl~[acp] reductase classified, for example, under EC 1 , 1.1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%:·, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3-dehydroundecanedIoyl-[acp] methyl ester by a polypeptide having the activity of a 3-hydroxyacyl~[acp] dehydratase classified, for example, under EC 4.2, 1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to undecanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl-facpj reductase classified, for example, under EC 1.3.1 .10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Using NABPH-Spedfk Enzymes to Produce Undecanedioyl-CoA Methyl Ester as a Centra! Precursor Leading to Cn Building Blocks

In some embodiments, undeeanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via four cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to nonanedioyl~CoA methyl ester as described above; followed by conversion to 3~oxo-imdecanedioyl~CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1 ,41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2,3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-utidecane4ioyl-CoA methyl ester by a polypeptide having the activity of a 3-oxoacyl~ [acpj reductase classified, for example, under EC 1.1 .1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3-hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1.1.1.157 (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4), or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC 1.1.1.36 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydroimdecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1.1 19 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 48); followed by conversion to undecanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyl~[acp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 6) or a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.38 or EC 1.3,1 ,8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7). Pathways Using ADH-Specifk Enzymes to Produce UradecaRedioyl-CoA Methyl Ester as a Centra! Precursor Leading to Cu Building Blocks

in some embodiments, undecanedioyl-CoA methyl ester is synthesized from the centra! metabolite propanedioyl-CoA. via four cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to nonanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo~undeeanedioyl~CoA methyl ester by a polypeptide having the activity of a fi~keioacyi~[acp] synthase classified, for example, under EC 2,3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothiolase classified, for example, under EC 2.3,1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence, of SEQ ID NO: 17); followed by conversion to 3~ hydroxy-undecanedioyl-CoA methyl ester by a polypeptide having the activity of a 3- h droxyacyl-CoA dehydrogenase classified, for example, under EC 1 , 1.1.35 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3- dehydroundecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl- CoA hydratase classified, for example, under EC 4.2.1.17 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to undecanedioyl-CoA methyl ester by a polypeptide having the activit of a tram-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 1 15) or a polypeptide having the activity of a enoyl-[acp] reductase classified, for example, under EC 1.3.1,9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 46). Pathways Usin Undeeanedioyl-CoA Methyl Ester or L¾decaKedioyl~facp] Methyl Ester as Central Precursors to Undeeanediok Add

in some embodiments, undecanedioic acid is synthesized from the central precursor, undecanedioyl-[aep] methyl ester, by conversion of undecanedioyl~[aep] methyl ester to monomethyl undecanedioate by a polypeptide having the activity of a thioesierase classified, for example, under EC 3.1.1.2, EC 3.1.1.5, or EC 3.1.2.-, such as EC 3.1.2, 14, EC 3.1.2.21, or EC 3.1.2.27. or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to undecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, undecanedioic acid is synthesized from the centra! precursor, undecanedioyl-CoA methyl ester, by conversion of undecanedioyl-CoA methyl ester to monomethyl undecanedioate by a polypeptide having the activity of a thioesierase classified, for example, under EC 3.1.1.2, EC 3,1 ,1.5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2.21, or EC 3.1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to undecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 ).

In some embodiments, undeeanedioie acid is synthesized from the central precursor, undecanedioyl-CoA methyl ester, by conversion of undecanedioyl-CoA methyl ester to monomethyl undecanedioate by a polypeptide having the activity of a CoA-transferase such as a glutaconate Co A-ir cms f erase classified, for example, under EC 2.8.3.12 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-ligase classified, for example, under EC 6,2.1 ,5; followed by conversion to undecanedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, undecanedioic acid is synthesized from the central precursor, imdecanedioyi-CoA methyl ester, by conversion of undecanedioyl-CoA methyl ester to methyl 11-oxoundeeanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, under EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19. a polypeptide having the activity of a succinate semiaidehyde dehydrogenase classified, for example, under EC 1.2.1 ,76. or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2,1.52; followed by conversion to monomethyl undecanedioate by a polypeptide having the activity of a non-acylating NAD- dependent aldehyde dehydrogenase (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1 ), a 7-oxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SJ!iQ ID NO: 13), a 6- oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g.. a polypeptide having at least 50%, at least 60%». at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 1 ), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to undecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, a least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, undecanedioic acid is synthesized from the central precursor, methyl 11-oxoundecanoate, by conversion of methyl 1 1 -oxoimdecanoate to monomethyl undecanedioate by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7- oxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%», at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to imdecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). Pathways Using UndecsHedioyi-CoA Methyl Ester or Methyl 11-Oxoisndecanoate as a Central Precursor to il-Amhiouftdecanoate

In some embodiments, 11-aminoundecanoate is synthesized from the central precursor. undecanedioyl-CoA methyl ester, by conversion of undecanedioyl-CoA methyl ester to methyl 11-oxoundecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semiald hyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 1 1 -oxoundecanoate to monomethyS 11-aminoundecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6,1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 : followed by conversion of monomethyl 1 1 -aminoundecanoate to 1 1 - aminoundecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 1 1 -amino undecanoate is synthesized from the central precursor, methyl 11-oxo undecanoate, by conversion of methyl 1 1 -oxoundecanoate to monomethyl 11-aminoundecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1 .18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 1 l-ammoundecanoate to 11- aminoundecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 1 1-aminoundecanoate is synthesized from the central precursor, monomethyl undecanedioate, by conversion of monomethyl undecanedioate to methyl 1 1 -oxoundecanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphop ntetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%», or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of methyl 1 1 -oxoundecanoate to monomethyl 1 1-aminoundecanoate by a polypeptide having the activity of a aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6,1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5.4,3.8, or a polypepiide having at least 50%, at least 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 1 1- aniinoundeeanoate to 1 1-aminoundecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 1 1-aminoundecanoate is synthesized from the central precursor, undecaiiedioic acid, by conversion of undecaiiedioic acid to 1 1-oxoundeeanoate by a polypeptide having the activity of a carboxyl te reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheim transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 1 1 - oxoundecanoate to l l-aminoundecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2,6.1 ,11, EC 2.6.1.13, EC 2,6.1.18, EC 2.6.1.19, EC 2.6,1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

Pathways Using il-A inosmdecaaoate, ll~Hydr xyimdecanoate, or 11- Oxowndeeanoate as Central Precursors to U¾decaKe-l Jl-Diamiiie

In some embodiments, undecane-1 ,1 1 -diamine is synthesized from the central precursor, 11 -amino undeeanoate, by conversion of l l-aminoundecanoate to 1 1- aminoundecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 1 1 -aminoundecanal to undecane- 1,11 -diamine by a polypeptide having the activity of an aminotransferase classified, for example, under E EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 1 16- 138 or SEQ ID NOs: 167-181.

The carhoxylate reductase encoded by the gene product of car and the phosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunetional C4 and C5 earboxylic acids (Venkitasiibramanian et al, Enzyme and Microbial Technology, 2008, 42, 130 - 137).

In some embodiments, undecane- 1,1 1 -diamine is synthesized from the central precursor, 1. 1 -hydroxyundecanoate. by conversion of 1 1-hydroxvuiidecanoate to 1 1- hydroxyiuidecanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at leas 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least. 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 1 1 -hydroxyundecanal to 1 1 -aminoundeeanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6, 1.11, EC 2.6.1.13, EC 2.6,1 .18, EC 2,6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to 11 -aminoundecanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21, EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to undecane-1 ,1 1- diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6, 1.19, EC 2.6.1 ,29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

In some embodiments, undecane- 1,11 -diamine is synthesized from the central precursor, 1 1-aminoundecanoate, b conversion of 1 1 -amino ndecanoate to Nl l~acetyl-l l- aminoundecanoate by a polypeptide having the activity of an N~ cetyltr nsferase such as a lysine N-acetyltransferase classified, for example, under EC 2.3.1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to Nl 1 -acetyl- 1 i- aminoundecanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1 ,2,99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to NI l-acetyl-1,1 1-diaminoundecane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1 ,-, such as EC 2.6.1 .1 1. EC 2,6.1.13. EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%o, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to undecane-1 ,11 -diamine by a polypeptide having the activity of an acetylputrescine de cylase classified, for example, under EC 3.5.1 ,62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, undecane-1,11 -diamine is synthesized from the central precursor, 1 1 -oxoundecanoate, by conversion of l l-oxoundecanoate to 1,1 1-undeeanedial by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the acti vity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion to 11- aminoundecanal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1 , 13, EC 2.6.1.18, EC 2,6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1 ,64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by- con version to undecane-1,1 1 -diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6.1.11, EC 2.6,1 , 13, EC 2,6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6, 1 .82, EC 4.1.1 ,64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 .

Pathwa s Using UBdeeanedioic Acid or UKdecanedioyl-CoA Methyl Ester as a Cen ral Precursor to ll-iiydroxyimdeeaijoaie

In some embodiments, l l-hydroxyundecanoaie is synthesized from the central precursor, undecanedioic acid, by conversion of undecanedioic acid to 1 1 -oxoundeeanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99,6 or a polypeptide having at least 50%), at least 60%, at least 70%), or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 11- hydroxyundecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1 ,1.1.- such as a 6-hydroxykexanoate dehydrogenase classified, for example, under EC 1.1.1.258 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoaie dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, ai least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a polypeptide having the activity of a 4-hydroxyhuiyrate dehydrogenase (e.g. _. , a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23).

In some embodiments, 11 -hydroxyundecanoate is synthesized from the central precursor, undecanedioyl-CoA methyl ester, by conversion of undeeanedioyl-CoA methyl ester to methyl 11-oxoundeeanoate by a polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least. 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdekyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 1 l-oxoundeeanoate to 11 -oxoundecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%, or a least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 1 1 -hydroxyundecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1,1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase classified, for example, under EC 1 , 1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23).

Pathways Using ll-H drox nndecanoaie as a Central Pre€¾srsor to Lli-Un ecsmedioi

In some embodiments, 1 ,1 1-undeeanediol is synthesized from the central precursor, 1 l-hydroxyirndecanoate, by conversion of l l-hydroxyundeeanoate to 1 1 -hydroxyundeeanal by a polypeptide having the activity of a carboxylaie reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteth ine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 11- hydroxyundecanal to 1,11-undecanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 .1. L- (e.g.. EC 1.1.1.1. EC 1.1.1.2, EC 1 .1.1.21 , EC 1.1.1 ,61 , or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Pathways Using 1.1 -Amiisousidecassoate as a Central Precursor to ll-Amh et rideeanol

In some embodiments, l l-aniinoundecanol is synthesized from the central precursor, 11-aminoundecanoate, by conversion of i l~aminoundecanoate to 1 1 -aminoundeeanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 1 1- aminoundecanal to 1 1-aminoundecanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC E L 1.1, EC 1 .1.1.2, EC 1.1.1.21, EC 1.1.1.61, or EC 1 ,1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23). Co Biochemical Pathways

Pathways Vmng NAOFH-Specific Enzymes to Produce TridecaBedioyl~[acp] Methyl Ester as a Central Precursor Leading to Cn Bmildiog Blocks

in some embodiments, tridecanedioyl-[acp] methyl ester is synthesized from the central metabolite propanedioyl- aep] via five cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-[acp] to undecanedioyi-j aep] methyl ester as described above; followed by conversion to 3~oxo-trid.ecanedioyl-[acp] methyl ester by a polypeptide having the activity of a fi-keioacyl-facp] synthase classified, for example, under EC 2.3,1.- (e.g., EC 2.3.1.41 or EC 23.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least. 7Q%>, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16): followed by conversion to 3-hydroxy-tridecanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3-oxoacyl~[acp] reductase classified, for example, under EC 1.1 .1 ,100 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); .followed by conversion to 2,3-dehydrotriclecanedioyl-[aep] methyl ester by a polypeptide having the activity of a 3 '-hydroxyacyi-facp] ' dehydratase classified, for example, under EC 4.2.1 ,59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to tridecanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoy!-facp] reductase classified, for example, under EC 1.3.1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Using NADPH-Speeifk Enzymes to Produce Trideca&edioyl-CoA Methyl Ester as a Central Precursor .Leading to C B iidisig Blocks

In some embodiments, tridecanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via five cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to undecanedioyl-CoA methyl ester as described above: followed by conversion to 3-oxo-tridecanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2,3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2.3.1 , 16 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydrox -tridecanedioyl~CoA methyl ester by a polypeptide having the activity of a 3-oxoacyl- [acp] reductase classified, for example, under EC 1.1. LI 00 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3~hydraxyacyl-CoA dehydrogenase classified, for example, under EC 1.1.1.157 (e.g., a polypeptide having at least. 50%, at least 60%>, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4), or a polypeptide having the activity of an acetoaceiyl-CoA reductase classified, for example, under EC 1.1.1.36 (e.g.. a polypeptide having at least 50%, at least 60%. at least 70%, or at least. 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydrotrideeanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydrolase classified, for example, under EC 4.2.1.119 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to tridecanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyl-facp] reductase classified, for example, under EC 1.3.1 .10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6) or a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.38 or EC 1.3.1 ,8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 114 or SEQ ID NO; 1.15).

Pathways Using NADH-Speeifk Enzymes io Produce Tridecanedioyl-CoA Methyl Ester as a Central Precursor Leading to C Building Blocks

In some embodiments, trideeanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyi-CoA via five cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to undeeanedioyl~CoA methyl ester as described above; followed by conversion to 3-oxo-tridecanedioyl-CoA methyl ester by a polypeptide having the activity of a j]~ketoacyl~[acp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a fi-fatothiolase classified, for example, under EC 2.3. ί .16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-tridecanedioyl-CoA methyl ester by a polypeptide having the activity of a i- hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1.1.1.35 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3- dehydrotridecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydrolase classified, for example, under EC 4.2.1.17 (e.g., e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to tridecanedioyl-CoA methyl ester by a polypeptide having the activity of a tran$~2~enoyl-CoA reductase classified, for example, under EC 1.3.1 ,44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 1 15) or a polypeptide having the activity of a enoyl-facp] reductase classified, for example, under EC 1.3.1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46).

Pathways Using Tridecanedioy!-CoA Methyl Ester or Tridecanedioyl-[acp] Methyl Ester as Central Precursors to Tridecanedioic Acid

in some embodiments, iridecanedioic acid is synthesized from the central precursor, tridecanedioyl~[acp] methyl ester, by conversion of trideeanedioyl-j acp] methyl ester to raonomethyl tridecanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1.1.2, EC 3.1.1.5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2,21, or EC 3.1.2,27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to iridecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

in some embodiments, iridecanedioic acid is synthesized from the central precursor, tridecanedioyl-CoA methyl ester, by conversion of tridecanedioyl-CoA methyl ester to n onomethyl tridecanedioate by a polypeptide having die activity of a thioesterase classified, for example, under EC 3.1.1.2, EC 3.1.1.5, or EC 3.1.2.-, such as EC 3.1.2, 14, EC 3.1.2,21, or EC 3.1.2,27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to tridecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, tridecanedioic acid is synthesized from the central precursor, tridecanedioyi-CoA methyl ester, by conversion of tridecanedioyl-CoA methyl ester to monomethyl tridecanedioate by a polypeptide having the activity of a CoA-transferase such as a glutaconate CoA-transferase classified, for example, under EC 2.8.3.12 (e.g., a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-ligase classified, for example, under EC 6,2.1.5; followed by conversion to tridecanedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3,1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, tridecanedioic acid is synthesized from the central precursor, tridecanedioyl-CoA methyl ester, by conversion of tridecanedioyi-CoA methyl ester to methyl 13-oxotridecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, under EC 1.2,1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 8), a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19. a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1 ,2.1.52; followed by conversion to monomethyl tridecanedioate by a polypeptide having the activity of a non~acylating NAD- dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 11), a 7-oxoheptanoate dehydrogenase classified, for example, under EC 1.2.L- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ Π) NO: 13), a 6- oxohexanoate dehydrogenase classified, for example, under EC 1 ,2, 1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%s sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1 ,2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to tridecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51), In some embodiments, tridecanedioic acid is synthesized from the central precursor, methyl 13~oxotrideeanoate, by conversion of methyl 13~oxotridecanoate to monomethyl tridecanedioate by a polypeptide having the activity of a non-acylaiing NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7~ oxohepianoate dehydrogenase classified, for example, under EC 1.2.L- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6~oxohexanoate dehydrogenase classified, for example, under EC 1.2.1 ,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to tridecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

Pathways Using Tridecaiiedioyi-CoA Methyl Ester or Methyl 13-Oxotridecas¾oate as a Central Precursor to 13~Ammotrideeanoate

In some embodiments, 13-aminotridecanoa.te is synthesized from the central precursor, tridecanedioyl-CoA methyl ester, by conversion of tridecanedioyl-CoA methyl ester to methyl 13~oxotridecanoate by a polypeptide having the activity of an acety!ating aldehyde dehydrogenase classified, for example, EC 1 ,2.1, 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semiaidehyde dehydrogenase classified, for example, under EC 1.2.1 ,76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 13~oxotrideeanoate to monomethyi 13-aminotridecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1.-, such as EC 2,6.1.11, EC 2.6.1.13, EC 2,6.1.18, EC 2.6.1 ,19, EC 2.6, 1.29, EC 2.6.1 ,48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs; 167-181 ; followed by conversion of monomethyi 13~aminotridecanoate to 13- aminotridecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ D NO: 50 or SEQ ID NO: 51).

in some embodiments, 13 -aminotridecanoate is synthesized from the central precursor, methyl 13 -oxotridecanoate, by conversion of methyl 13 -oxotridecanoate to monomethyi 13- aminotridecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2,6, 1.18, EC 2.6.1.19, EC 2.6.1 ,29, EC 2.6.1.48, or EC 2.6,1 .82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyi 13 -aminotridecanoate to 13-aminotridecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 13-aminotridecanoate is synthesized from the central precursor, monomethyi tridecanedioate, by conversion of monomethyi tridecanedioate to methyl 13- oxotrideeanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of methyl 13~oxotridecanoate to monomethyl 13-arninotrideeanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1 , 1.64, or EC 5,4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 13- ammotridecanoate to 13 -ammotridecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

in some embodiments, 13-aminotridecanoate is synthesized from the central precursor, trideeanedioie acid, by conversion of tridecanedioie acid to 13~oxotridecanoate by a polypeptide having the activity of a. carhoxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ⁾ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 13- oxotridecanoate to 1.3 -ammotridecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6,1.11, EC 2.6, 1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1 ,29, EC 2.6.1.48, or EC 2.6, 1.82, EC 4.1.1 ,64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167481.

Pathways Using 13-Am otrideca3iea†e, 13-Hydrox trideeaaoate, or 13-Oxotridecanoate Ceotral Precursors to Tridecaiie-l, ~DiaEiiiie

In some embodiments, tridecane- 1,13 -diamine is synthesized from the central precursor, 13-aminotridecanoate, by conversion of 13 -ammotridecanoate to 13- arninotridecanal by a polypeptide having the activity of a c rboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanletheine transferase classified, for example, under EC 2,7.8.~, such as 2,7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by ^¬ conversion of 13-aminotridecanal to tridecane- 1 ,13 -diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2,6.1 , 13, EC 2.6.1.18, EC 2.6, 1 , 19, EC 2.6, 1.29, EC 2.6,1.48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%), at least 60%*, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

The carboxylate reductase encoded by the gene product of car and the pkosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunctional C4 and C5 carboxylie acids (Venkitasubramanian et /., Enzyme and Microbial Technology, 2008, 42, 130 ^■■■■ 137).

In some embodiments, tridecane- 1, 13 -diamine is synthesized from the central precursor, 13-hydroxytridecan.oate, by conversion of 13-hydroxytridecanoate to 13- hydroxyiridecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 .2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a pkosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least. 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 13 -hydroxyiridecanal to 13-aminotrideeanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2,6.1.18, EC 2.6,1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to 13-aminotridecanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g.. EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1 .21 , EC 1.1 .1.61, or EC LI .1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to tridecane~lJ 3~ diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1,29, EC 2.6.1.48, or EC 2,6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

In some embodiments, ttidecane-l ,L3 -diamine is synthesized from the central precursor, 13-aminotridecanoate, by conversion of 13-aminotridecanoate to N13-acetyl-13- aminotridecanoate by a polypeptide having the activity of an N-ace!yltransferase such as a lysine N-aceiyltransferase classified, for example, under EC 2,3.1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to Nl 3 -acetyl- 13- aminotridecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%), at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least. 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to Nl 3-acetyl- 1. , 13-diarainotridecane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2,6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2,6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1,82, EC 4,1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs; 167-181 ; followed by conversion to tridecane- 1,13 -diamine by a polypeptide having the activity of an acetylputrescine deacylase classified, for example, under EC 3.5.1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, tridecane- 1,13 -diamine Is synthesized from die central precursor, 13-oxotridecanoate, by conversion of 13-oxotridecanoate to 1 ,13-tridecanedial by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 13- aminotridecanal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6.1 .82, EC 4,1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-1 1 ; followed by conversion to tridecane- 1 , 13 -diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1 , 11 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1 ,19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6.1.82, EC 4.1 .1.64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

Pathways Using Tridecanedioie Acid or Tridecanedioyi-CoA Methyl Ester as a Central Precursor to 13~Hydroxytridecanoate

In some embodiments, 13-hydroxytridecanoate is synthesized from the central precursor, tridecanedioic acid, by conversion of tridecanedioic acid to 13-oxotrideeanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 13- hydroxytridecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1,1.- such as a 6~hydroxyhexanoaie dehydrogenase classified, for example, under EC L i .1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypent noate dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a polypeptide having the activity of a 4-hydroxyb tyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 23).

In some embodiments, 13 -hydroxytridecanoate is synthesized from the central precursor, tridecanedioyl-CoA methyl ester, by conversion of tridecanedioyl-CoA methyl ester to methyl 13-oxotridecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2,1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1.8), a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1 ,2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 13-oxotridecanoate to 13-oxotridecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 13 -hydroxytridecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1 , 1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1 , 1.1 ,258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%,· or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 8), a polypeptide having the activity of a 5-hydroxypentanoale dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at feast 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 21 ), or a polypeptide having the activity of a 4-hydroxybuiyraie dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of SEQ ID NO: 23).

Pathways Using 13-Hydrosytridec¾ii ste as a C ntr l Precursor to 1,13-TrideeaBedioI

In some embodiments, 1 , 13-tridecanediol is synthesized from the central precursor, 13-liydroxytridecanoate, by conversion of 13-hydroxytridecanoate to 13-hydroxytridecanai by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%?, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteikeine transferase classified, for example, under EC 2,7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%t sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 13- hydroxyiridecanal to 1,13-tridecanediol by a polypeptide having die activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1 , EC 1.1.1 ,2, EC 1.1.1.21 , EC 1 , 1.1.61 , or EC 1 , 1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Pathways Usiag 13~Aminotr ecsmoate m a Central Precursor to 13-Aminotrideean l

In some embodiments, 13-aminotridecanol is synthesized from the central precursor, 13-aminotridecanoate, by conversion of 13 -amino trideeanoate to 13-aminotridecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ IP NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8,-, such as 2.7,8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ D NOs: 54-57); followed by conversion of 13- aminotridecanal to 13-aminotridecanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 ,1 ,1.- (e.g., EC 1.1.1.1 , EC 1.1 , 1 ,2, EC 1 ,1 , 1.21, EC 1.1.1 ,61. or EC 1.1 ,1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Ci Biochemical Pathways

Pathways Using NADPH-Specific Enzymes to Produce Pentadecaaedioyi~[acp] Methyl Ester as a Centra! Precursor Leading to€. \$ Building Blocks

In some embodiments, pentadecanedioyl-[acp] methyl ester is synthesized from the central metabolite p.ropanedioyi-[acp] via six cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-jacp] to tridecanedioyl-[acp] methyl ester as described above; followed by conversion to 3-oxo~pentadecanedioyl~[aep] methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2,3.1.41 or EC 2.3,1 , 179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3-hydroxy~pentadecanedioyl~[aep] methyl ester by a polypeptide having the activity of a 3~oxoacyl-[acp] reductase classified, for example, under EC 1 , 1 , 1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3-dehydropentadecanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3~hydroxyacyl~[acp] dehydratase classified, for example, under EC 4.2,1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 1 ); followed by conversion to pentadecanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoy!-facp] reductase classified, ^" for example, under EC 1.3,1, 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%* sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Usin NAD PB -Specific Enz mes to Produce FeiitadecaKedioyi-CoA Methyl Ester as a Central Precursor Leading to Ci§ Building Blocks

In some embodiments, pentadecanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via six cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to tridecanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo-pentadecanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-f cp] synthase classified, for example, under EC 2.3.1." (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2.3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-pentadecanedioyl-CoA methyl ester by a polypeptide having the activity of a 3- oxoacy!-facp] reductase classified, for example, under EC 1.1.1.100 (e.g., a polypeptide having at least 50%;, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having die activity of a 3-hydroxyacyl- CoA dehydrogenase classified, for example, under EC 1 ,1.1.157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4), or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC 1,1.1.36 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2 ₅ 3-dehydropentadecanedioyl~CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydrolase classified, for example, under EC 4.2.1.119 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to pentadecanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 13. L- such as an enoyl-facp] reductase classified, for example, under EC 1.3.1 ,10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 6) or a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1.38 or EC 1 ,3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7).

Pathways Using NADH-Spedfk Enzymes to Produce Peiiiadecasiedioy!-CoA Methyl Ester as a Central Precursor Leading to C15 Building Blocks

In some embodiments, pentadecanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via six cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to tridecariedioyi-CoA methyl ester as described above; followed by conversion to 3-oxo-pentadeeanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facpj synthase classified, for example, under EC 2.3. L- (e.g., EC 2,3.1 ,41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothiolase classified, for example, under EC 2,3.1.16 (e.g., a polypeptide having at least 5Q%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-pentadecanedioyl-CoA methyl ester by a polypeptide having the activity of a 3- hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1.1.1.35 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3- dehydropentadecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl- CoA hydratase classified, for example, under EC 4.2.1.17 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to pentadecanedioyl-CoA methyl ester by a polypeptide having the acti vity of a irans-2-enoyl~CoA. reductase classified, for example, under EC 1.3.1.44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 1 15) or a polypeptide having the activity of a enoyl-facp] reductase classified, for example, under EC 1.3.1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at. least 85% sequence homology to the amino acid sequence of SEQ ID NO; 46).

Pathways Using Peiii¾decanedi yl-C A Methyl Ester or Peniaclecanedioyl-faep] Methyl Ester as Central Precursors to Pesitade ne ioie Add

In some embodiments, pentadecanedioic acid is synthesized from the central precursor, pentadecanedioyl-[acp] methyl ester, by conversion of pentadeeanedioyl~[aep] meihyi ester to monomethyi pentadecanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1 ,1.2, EC 3.1.1 ,5, or EC 3.1.2,-, such as EC 3.1.2.14, EC 3.1.2,21 , or EC 3.1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to pentadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

in some embodiments, pentadecanedioic acid is synthesized from the central precursor, pentadecanedioyi-CoA methyl ester, by conversion of pentadecanedioyi-CoA methyl ester to monomethyi pentadecanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3,1.1.2, EC 3.1.1.5. or EC 3.1.2,-, such as EC 3.1.2.14, EC 3.1.2,21, or EC 3, 1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to pentadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

in some embodiments, pentadecanedioic acid is synthesized from the central precursor, pentadecanedioyi-CoA methyl ester, by conversion of pentadecanedioyl-CoA methyl ester to monomethyi pentadecanedioate by a polypeptide having the activity of a CoA- transferase such as a ghitaeonate CoA-tr nsferase classified, for example, under EC 2.8.3,12 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence bomology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-Iigase classified, for example, under EC 6.2.1.5; followed by conversion to pentadecanedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO; 51).

in some embodiments, pentadecanedioic acid is synthesized from the central precursor, pentadecanedioyl-CoA methyl ester, by conversion of pentadecanedioyl-CoA methyl ester to methyl 15-oxopentadecanoate by a polypeptide having the activity of an acelylating aldehyde dehydrogenase classified, for example, under EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19. a polypeptide having the activity of a succinate semialdekyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutaraie dehydrogenase classified, for example, under EC 1.2.1 ,52; followed by conversion to monomethyl pentadecanedioate by a polypeptide having the activity of a non-acylating NAD-ckpendent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%» sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7-Qxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at. least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.13 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%», or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to pentadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, pentadecanedioic acid is synthesized from the central precursor, methyl 15-oxopentadeeanoate, by conversion of meth l I S-oxopentadecanoate to monomethyl pentadecanedioate by a polypeptide having the activity of a non-acylating NAD- dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 11), a 7-oxoheptanoate dehydrogenase classified, tor example, under EC 1.2.L- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6~ oxohexanoaie dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1 ,2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to pentadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

Pathways Using Pentadeeanedioyl-CoA Methyl Ester or Methyl IS-Oxopesitadecasioate as a Central Precursor to 15-Aminopentadecanoate

in some embodiments, 15-aminopentadecanoate is synthesized from the central precursor, pentadeeanedioyl-CoA methyl ester, by conversion of pentadeeanedioyl-CoA methyl ester to methyl 15-oxopentadecanoate by a polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1 ,76, or a polypeptide having the activity of an oxogkdarate dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 15-oxopentadecanoate to monomethyl 15-aminopentadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1.64, or EC 5,4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 15-aminopentadecanoate to 15- aminopentadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 50 or SEQ ID NO: 51).

in some embodiments, 15-aminopentadecanoate is synthesized from the central precursor, methyl 15-oxopentadecanoate, by conversion of methyl 15-oxopentadecanoate to monomethyl 15-aminopentadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6,1 .18, EC 2,6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 15-aminopentadecanoate to 15- aminopentadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO; 51).

In some embodiments, 15-aminopentadecanoate is synthesized from the central precursor, monomethyl pentadecanedioate, by conversion of monomethyl pentadecansdioate to methyl 15-oxopentadecanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 In combination with a polypeptide

1 having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54- 57); followed by conversion of methyl 15 -oxopentadecanoate to monomethyl 15- aminopentadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2,6.1.13, EC 2.6.1 ,18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181; followed by conversion of monomethyl IS-aminopentadecanoate to 15-aminopentadeeanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%. at least 60%!, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 15-aminopentadecanoate is synthesized from the central precursor, pentadecanedioic acid, by conversion of pentadecanedioic acid to 15- oxopentadecanoate by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion of 15~oxopentadecanoate to 15-aminopentadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6, 1.1 1, EC 2.6, 1.13, EC 2.6.1.18, EC 2,6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1 , 1 .64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181. Pathways Using IS-Ammope adeeanoate, IS-Hydroxypejitadeearsoate, or 15- OxopeniadeeaEoafe as Central Precursors to Pc!itadecaue-l,i§-J>ksHH*£

in some embodiments, pentadecane- 1 , 15-diamine is synthesized from the central precursor, 15-aminopentadecanoate, by conversion of 15-aminopentadecanoate io 15- aminopentadecanal by a polypeptide having the activity of a carhox l te reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion of 15-aminopentadecanal to pentadecane- 1 , 15-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2,6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16- 138 or SEQ ID NOs: 167-181 ,

The carhoxylate reductase encoded by the gene product of car and the phosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunctional C4 and C5 carboxylic acids (Yenkitasubramanian et al, Enzyme and Microbial Technology, 2008, 42, 130 - 137).

in some embodiments, pentadecane-!, 15-diamine is synthesized from the central precursor, 15-hydroxypentadecanoate, by conversion of 15-hydroxypentadecanoate to 15- hydroxypentadecanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs; 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); fol lowed by conversion of 15-hydroxypentadeeanal to 15-aminopentadecanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1 ,13, EC 2.6.1.18, EC 2,6.1.19, EC 2.6.1.29, EC 2,6.1.48, or EC 2.6.1.82, EC 4.1 ,1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116- 138 or SEQ ID NOs: 167-181 ; followed by conversion to 15-aminopentadeeanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21, EC 1.1.1.61 , or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); foilowed by conversion to pentadecane~l ,15~di amine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1.-, such as EC 2.6.1.11 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

in some embodiments, pentadecane~l,15~diamine is synthesized from the central precursor, 15-aminopentadecanoate, by conversion of 15-aminopentadecanoate to N15~aeetyl~ 15-arninopentadeeanoate by a polypeptide having the activity of an N-acetyltramferase such as a lysine N-acetyltransferase classified, for example, under EC 2.3.1.32 (e.g., a polypeptide, having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to N15-acetyi-15- aminopentadecanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion to Nl 5-acetyl- 1 , 15-diaminopentadecane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6.1.1 1 , EC 2.6.1 .13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1 .82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to pentadeeane- 1,15 -diamine by a polypeptide having the activity of an acetylpidrescme deacylase classified, for example, under EC 3.5,1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, pentadeeane- 1 ,15-diamine is synthesized from the central precursor, 15~oxopentadeeanoate, by conversion of 15-oxopentadecanoate to 1 ,15- pentadeeanedial by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphop ntetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%>, at least 70%>, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 15-aminopentadeca.nal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1 ,11, EC 2,6.1.13, EC 2.6.1 ,1 8, EC 2.6.1 ,19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6.1 ,82, EC 4.1 ,1 .64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%», or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to pentadecane- 1 , 15~diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6,1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1 ,1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 , Pathways Using PeBtadeeaaedioie Add or PesstadeeaEedioyl-CoA Methyl Ester as a Cen ral Precursor to IS-Hydroxyperaiadecassoate

In some embodiments, 15-hydroxypentadeeanoate is synthesized from the central precursor, pentadecanedioic acid, by conversion of pentadecanedioic acid to 15- oxopentadecanoate by a polypeptide having the activity of a carboxylase reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85%) sequence homology to the amino acid sequence of any one of SEQ ID NOs; 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanietheine transferase classified, for example, under EC 2,7.8.-, such as 2.7,8,7 (e.g., a polypeptide having at least 50%s, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 15-hydroxypentadecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1. 1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1 .258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%s, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO; 8), a polypeptide having the activity of a S hydraxypentanoaie dehydrogenase classified, for example, under EC 1.1.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a poiypeptide having the activity of a 4-hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 23).

In some embodiments, 15-hydroxypentadeeanoate is synthesized from the central precursor, pentadeeanedioyl~CoA methyl ester, by conversion of pentadecanedioyl-CoA methyl ester to methyl 15-oxopentadecanoate by a polypeptide having the activity of an acefylating aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%!, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%o sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1 ,2.1 .76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1 ,2.1.52; followed by conversion of methyl 15-oxopentadecanoate to 15-oxopentadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ΪΪ) NO: 50 or SEQ ID NO: 51); followed by conversion to 15-hydroxypentadeeanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1 ,258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%;, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a S-hydroxypentanoate dehydrogenase classified, for example, under EC 1.1.1 ,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxyb tyraie dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 23).

Pathways Usmg IS-IIydroxypeBtadecanoaie as a Centra! Precursor to 1,15- Pentadeeaiiediol

In some embodiments, 1,15-pentadecanediol is synthesized from the central precursor, 15-hydroxypentadecanoate, by conversion of 15-hydroxypentadecanoate to 15- hydroxypentadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SKQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%» sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 15-hydroxypentadecanal to 1 ,15-pentadecanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1 ,1 , 1.1, EC 1 .1.1 ,2, EC 1.1 ,1.21 , EC 1 ,1.1.61, or EC 1.1.1 ,1 84) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Pathways Using !5~Amim>pen a eeaKoate as a Central Precursor to 15- Ammopentadeeano!

In some embodiments, 15-aminopentadecanoS is synthesized from the central precursor, 15-aminopentadecanoate, by conversion of 15-aminopentadecanoate to 15- aminopentadecanal by a polypeptide having the activity of a carhoxy!ate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8,-, suc as 2,7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 15-aminopeniadecanal to 15-aminopentadecanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1 ,1.- (e.g., EC 1.1 .1.1, EC 1.1.1.2, EC 1.1.1 .21 , EC 1.1 .1.61, or EC 1 .1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 20-23).

C37 Biochemical Pathways

Pathways Usmg NADFH-Spectt!e Eraymes to Produce OeptadecaKcdioyl-[acp] Methyl Ester as ¾ Centra! Precursor Leading to Cn Building Blocks

In some embodiments, heptadecanedioyl-[aep] methyl ester is synthesized from the central metabolite propanedioyl-[acp] via seven cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-jacp] to pentadecanedioyl~[acp] methyl ester as described above; followed by conversion to 3-oxo-heptadeeanedioyl-[acp] methyl ester by a polypeptide having the activity of a fi~ toacyi~[acp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2,3.1.41 or EC 2.3.1 ,179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3~hydroxy~heptadeeanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3-oxoacyI-facpj reductase classified, for example, under EC LI , 1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3-dehydroheptadecanedioyl~[acp] methyl ester by a polypeptide having the activity of a 3~hydroxyacyl~[acp] dehydratase classified, for example, under EC 4.2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to heptadeeanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl~[acp] reductase classified, for example, under EC 13.1 ,10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Using NADPH-Specsfk Eii&ymes to Produce Heptssdec¾uedioyl~CoA Methyl Ester as a Central Precursor Leading to Ci ₇ Building Blocks

In some embodiments, heptadecanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via seven cycles of methyl-ester shielded carbon chain elongation by conversion, of propanedioyl-CoA to pentadeeanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo~heptadecanedioyl-CoA methyl ester by a polypeptide having the activity of a β-ketoacyl-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1 .179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2.3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-heptadecanedioyl-CoA methyl ester by a polypeptide having the activity of a 3- oxoacyl-facpj reductase classified, for example, under EC 1.1.1.100 (e.g., a poiypepiide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 5), a polypeptide having the activity of a 3-hydrox acyl- CoA dehydrogenase classified, for example, under EC 1 .1.1.157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 4), or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC L I .1,36 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 3); followed by conversion to 2,3-dehydroheptadecanedioyI-CoA methyl ester by a polypeptide ha ving the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2.1 ,119 (e.g., a polypeptide having at least 50%, at least 60%, at. least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to heptadecanedioyi-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3,1.- such as an enoyl-facp] reductase classified, for example, under EC 1.3, 1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6) or a trans-2~enoyl~CoA reductase classified, for example, under EC 1.3,1.38 or EC 1 ,3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%», or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7),

Pathways Using NADH-Specific Enzymes to Produce Hepiadecanedioyi-CoA Methyl Ester as a Central Free srsor Leading to Cn Biiilding Blocks

In some embodiments, heptadecanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via seven cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to pentadecanedioyl-CoA methyl, ester as described above; followed by conversion to 3-oxo-heptadecanedioyl-CoA methyl ester by a polypeptide having the activity of a β-keto cyl-f cp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2,3.1.41 or EC 2.3.1 , 179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothiolase classified, for example, under EC 2.3.1 ,16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-heptadecanedioyl-CoA methyl ester by a polypeptide having the activity of a 5- hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1 ,1.1.35 (e.g., a polypeptide having at least 50%>, at least 60%, at least 70%». or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2): followed by conversion to 2,3- dehydroheptadecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl- CoA hydratase classified, for example, under EC 4,2.1.1 7 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to heptadecanedioyl-CoA methyl ester by a polypeptide having the activity of a trans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1 ,44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 114 or SEQ ID NO: 115) or a polypeptide having the activity of a enoyl-facpj reductase classified, for example, under EC 1.3.1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 46).

Pathways Using Heptacleeaoedi y!-CoA Methyl Ester or Heptadecaiiedioyl-[acp] Methyl Ester as Ce tral Precursors to Ileptadecanedloie Acid

In some embodiments, heptadeeanedioic acid is synthesized from the central precursor, heptadecanedioyl>[acp] methyl ester, by conversion of heptadecanedioyl~[acp] methyl ester to monomethyl lieptadeeanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1.1.2, EC 3.1.1 ,5, or EC 3.1.2,-, such as EC 3.1.2.14, EC 3.1 ,2.21 , or EC 3.1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to heptadeeanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1.1 (e.g., a polypeptide having at least 50%, at least. 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, heptadeeanedioic acid is synthesized from the central precursor, heptadecanedioyl-CoA methyl ester, by conversion of heptadecanedioyl-CoA methyl ester to monomethyl lieptadeeanedioate by a polypeptide having the activity of a thioesterase classified, for example, under EC 3,1.1.2, EC 3, 1.1 ,5, or EC 3.1.2,-, such as EC 3.1 ,2, 14, EC 3.1 ,2.21, or EC 3, 1.2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-113 or SEQ ID NOs: 182-195; followed by conversion to heptadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1 ,1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, heptadecanedioic acid is synthesized from the central precursor, heptadecanedioyl-CoA methyl ester, by conversion of heptadecanedioyl-CoA methyl ester to monomethyl hepiadecanedioate by a polypeptide having the activity of a CoA- transferase such as a glutaconate CoA-tramferase classified, for example, under EC 2.8.3,12 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-!igase classified, for example, under EC 6.2.1,5; followed by conversion to heptadecanedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, heptadecanedioic acid is synthesized from the central precursor, heptadecanedioyl-CoA methyl ester, by conversion of heptadecanedioyl-CoA methyl ester to methyl 17-oxoheptadecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, under EC 1.2.1.10 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semi ldehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1 ,2,1.52; iollowed by conversion to monomethyl heptadecanedioate by a polypeptide having the activity of a non-acylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1 ), a 7~oxoheptanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohex noate dehydrogenase classified, for example, under EC 1.2.1.- (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to heptadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequenc of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, heptadecanedioic acid is synthesized from the central precursor, methyl 17-oxoheptadecanoate, by conversion of methyl 17-oxoheptadecanoaie to monomethyl heptadecanedioate by a polypeptide having the activity of a non~acylatlng NAD- dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 1), a 7-oxohepianoaie dehydrogenase classified, for example, under EC 1 ,2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6- oxohexanoate dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversio to heptadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at. [east 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51). Pathways Using Hepiadeearaedioy!-CoA Methyl Ester or Methyl 17-Oxoheptadecaiioate as 'Λ Centra! Precursor to IT-AmMoheptadecanoate

In some embodiments, 17-ammoheptadecanoate is synthesized from the central precursor, heptadecanedioyl-CoA methyl ester, by conversion of heptadecanedioyl-CoA methyl ester to methyl 17-oxoheptadecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1 ,2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semiaidehyde dehydrogenase classified, for example, under EC 1.2.1.76, or a polypeptide having the activity of an oxoglutarate dehydrogenase classified, for example, under EC 1.2.1 ,52; followed by conversion of methyl 17-oxoheptadecanoate to monomethyi 17-aminoheptadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6. LI 8, EC 2.6.1 .19, EC 2.6.1 .29, EC 2.6.1.48, or EC 2.6.1 .82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs; 167-181 ; followed by conversion of monomethyi 17-aminoheptadecanoate to 17- aminoheptadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, 17-arntn.oheptadecanoate is synthesized from the central precursor, methyl 17-oxoheptadeeanoate, by conversion of methyl 17-oxoheptadecanoate to monomethyi 17-aminoheptadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6, 1.11, EC 2.6.1 , 13, EC 2.6.1 , 18, EC 2,6.1.19, EC 2.6.1 ,29, EC 2,6.1.48, or EC 2.6.1.82, EC 4, 1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ D NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyi 17-aminoheptadecanoate to 17- aminoheptadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequenc homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51 ).

In some embodiments, 17-aminoheptadecanoate is synthesized from the central precursor, monomethyl heptadecanedioate, by conversion of monomethyl heptadecanedioate to methyl 17-oxoheptadeeanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 6Q%>, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54- 57); followed by conversion of methyl 17-oxoheptadecanoate to monomethyl 17- aminoheptadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4.1.1.64, or EC 5,4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least. 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 17-aminoheptadecanoate to 17-aminoheptadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 ,1 (e.g., a polypeptide having at least 50%, at least 60%), at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

In some embodiments, 1 7-aminoheptadecanoate is synthesized from the central precursor, heptadeeanedioic acid, by conversion of heptadecanedioic acid to 17- oxoheptadecanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-21 5 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequenc homology to the amino acid sequence of any one of SEQ ID NOs; 54-57); followed by conversion of 17~oxoheptadecanoate to 17~aminoheptadeeanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1 .18, EC 2.6.1,19, EC 2.6.1,29, EC 2.6.1.48, or EC 2.6.1 ,82, EC 4,1.1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

Pathways Using 1 -AmiKoheptadeeanoai j 17-Hydroxy epiadecasioate, or 17- Oxoheptedecanoaie as Central Precursors to Heptadeesme-l,17-Dkmine

In some embodiments, heptadecane-l ,17-diarnine is synthesized from the central precursor, 17-aminoheptadecanoate, by conversion of 17-aminoheptadecanoate to 17- aminoheptadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 17-aminoheptadecanal to heptadecane-l,17-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2,6.1.1 1 , EC 2,6.1.13, EC 2.6.1.18, EC 2,6.1.19, EC 2,6.1.29, EC 2.6,1.48, or EC 2,6.1.82, EC 4,1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16- 138 or SEQ ID NOs: 167-181.

The carboxylate reductase encoded by the gene product of car and the phosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunetional C4 and C5 carboxylic acids (Venkitasubramanian et al. Enzyme and Microbial Technology, 2008, 42, 130 - 137). In some embodiments, heptadecane~l ,17-diamine is synthesized from the central precursor, 17-hydroxyheptadecanoate, by conversion of 1 7-hydroxyheptadecanoate to 17- hydroxyheptadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs; 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 17-hydroxyheptadeeanal to 17-aminoheptadecanol by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11 , EC 2.6, 1.13, EC 2.6.1 , 18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5,4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16- 138 or SEQ ID NOs: 167-181 ; followed by conversion to 17-aminoheptadecanal by a polypeptide having the aciivity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1 , EC 1.1.1,2, EC 1.1.1.21, EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); followed by conversion to heptadecane-1 ,l 7~diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1 , EC 2.6.1.13, EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2.6, 1.48, or EC 2.6.1.82, EC 4.1 , 1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs; 167-181 ,

In some embodiments, heptadecane-1 , 17-diamine is synthesized from the central precursor, 17-aminohept.adecanoate, b conversion of 17-aminoheptadecanoate to N17-acetyl- I7-an inoheptadecanoate by a polypeptide having the activity of an N~acetyltransferase such as a lysine N-acetyltransfer se classified, for example, under EC 2.3, 1.32 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to N17-acetyl-17- a ninoheptadeeanal by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2.99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by ^¬ conversion to N17-acetyl-l ,17-diaminoheptadecane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6, 1.-, such as EC 2.6.1 , 1 1 , EC 2.6, 1.13, EC 2.6.1 , 18, EC 2,6.1.19, EC 2.6.1.29, EC 2.6.1 .48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to hepiadecane~l J 7~diamme by a polypeptide having the activity of an acelylputrescine deacylase classified, for example, under EC 3.5,1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, heptadecane-l,17-d amine is synthesized from the central precursor, 17-oxoheptadeeanoate, by conversion of 17~oxoheptadecanoate to 1 ,17- heptadecanediai by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or a least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 17-aminoheptadecanal by a polypeptide having th activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1.1 1, EC 2.6, 1.13, EC 2.6.1.18, EC 2.6.1 .19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6,1.82, EC 4.1.1 ,64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to heptadecane- 1 , 17-diamine by a polypeptide having the activity of an aminotransferase classified, for example, underEC 2.6,1,-, such as EC 2,6.1.1 1, EC 2.6,1.13, EC 2.6.1 .18, EC 2.6.1 .19, EC 2.6.1.29, EC 2.6.1.48, or EC 2,6.1.82, EC 4, 1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167481.

Pathways Using Heptadeeanedioie Aeid or Heptadeeaaedioyl-CoA Methyl Ester as a Central Precursor to 17-Hydroxyhepiadecauoaie

in some embodiments, 17~hydroxyhepiadecanoate is synthesized from the central precursor, heptadecanedioic acid, by conversion of heptadecanedioic acid to 17- oxohepiadecanoate by a polypeptide having the activity of a carhoxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 50%, at least 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7,8,-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed byconversion to 17-hydroxyhepiadecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-kydroxypenianoaie dehydrogenase classified, for example, under EC 1 , 1.1.- (e.g., a polypeptide having at least 50%, at least 6Q%>, at least 70%, or at least 85%o sequence homology to the amino acid sequence of SEQ YD NO: 21), or a polypeptide having the activity of a 4-hydroxyhun>rate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23). In some embodiments, 17-hydroxyheptadecanoate is synthesized from the central precursor, heptadeeanedioyl-CoA methyl ester, by conversion of heptadecanedioyl-CoA methyl ester to methyl 17-oxohepiadeeanoate by a polypeptide having the activity of an aceiylating aldehyde dehydrogenase classified, for example, EC 1.2,1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%o, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19. a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2.1 ,76, or a polypeptide having the activity of an oxoglutaraie dehydrogenase classified, for example, under EC 1.2.1.52; followed by conversion of methyl 17-oxoheptadecanoate to 17-oxoheptadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%s, at least 70%>, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 17-hydroxyheptadecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC I .I. I .- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5~hydroxypenianoate dehydrogenase classified, for example, under EC 1.1.1,- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptide having the activity of a 4-hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of SEQ ID NO: 23).

Pathways Using 17~Hydroxyfaeptsulec¾moate as a Central Precursor to 1,17- I-Iepfadeeanedie!

In some embodiments, l,17~heptadecanediol is synthesized from the central precursor, 17-hydroxyheptadecanoate, by conversion of 17-liydroxyheptadeeanoate to 1 - hydroxyheptadecanal by a polypeptide having the activity of a c rboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2,7.8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 17-hydroxyheptadecanal to I,17~heptadeeanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21, EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Pathways Using 17-AmiBdheptadeeanoate as a Central Precursor to 1 - Ammoheptadecamol

In some embodiments, 17-amlnoheptadeeanoi is synthesized from the central precursor. 1 -aminoheptadecanoaie, by conversion of 17-aminoheptadecanoate to 17- aminoheptadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs; 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine trans/erase classified, for example, under EC 2.7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 1 7-aminoheptadecanal to I 7~aminoheptadeeanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g., EC 1.1.1.1 , EC 1.1.1 .2, EC 1.1 .1.21, EC 1 , 1.1 ,61, or EC 1.1 ,1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23), Ci9 Biochemical Pathways

Pathways Using NADPH-Spediie Enzymes to Produce N©nadeca3ie i©yl~[aep] Methyl Ester M a Central Precursor Leading to Ci» Building Blocks

In some embodiments, nonadecanedioyl-[acp] methyl ester is synthesized from the central metabolite propanedioyl-[acp] via eight cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl~[acp] to heptadecanedioyl-[acp] methyl ester as described above; followed by conversion to 3-oxo~nonadecanedioyl-[acp] methyl ester by a polypeptide having the activity of a β-keloacyl-faep] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3.1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16); followed by conversion to 3~hydroxy~nonadecanedioyl~[acp] methyl ester by a polypeptide having the activity of a 3-oxoacyl-[acp] reductase classified, for example, under EC 1.1 .1.100 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at. least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5); followed by conversion to 2,3-dehydrononadecanedioyl-[acp] methyl ester by a polypeptide having the activity of a 3-hydroxyacyl~[acp] dehydratase classified, for example, under EC 4.2.1.59 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1); followed by conversion to nonadecanedioyl-[acp] methyl ester by a polypeptide having the activity of an enoyl~[acp] reductase classified, for example, under EC 1.3.1 , 10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 6).

Pathways Using NADPH-Spedfie Enzymes to Produce oiiadecanedioyl-CoA Meth l Ester as a Central Precursor Leading to C5 Building Blocks

In some embodiments, nonadeeanedioyl-CoA methyl ester is synthesized from the central metabolite propanedioyl-CoA via eight cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyl-CoA to heptadecanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo-nottadecanedioyi-CoA methyl ester by a polypeptide having the activity of a β-kefoacyi-facp] synthase classified, for example, under EC 2.3.1.- (e.g., EC 2.3,1.41 or EC 2.3.1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16), or a β-ketothiolase classified, for example, under EC 2.3.1 .16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-nonadecaiiedioyl-CoA methyl ester by a polypeptide having the activity of a 3- oxoacyl-facp] reductase classified, for example, under EC 1.1.1.100 (e.g., a polypeptide having at least. 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 5), a polypeptide having the activity of a 3-hydroxyacyl- CoA dehydrogenase classified, for example, under EC 1.1.1.157 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 4). or a polypeptide having the activity of an acetoacetyl-CoA reductase classified, for example, under EC 1 .1.1.36 (e.g., a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 3); followed by conversion to 2,3-dehydrononadecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl-CoA hydratase classified, for example, under EC 4.2,1.1 19 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 48); followed by conversion to nonadecanedioyl-CoA methyl ester by a polypeptide having the activity of a reductase classified, for example, under EC 1.3.1.- such as an enoyl~[acp] reductase classified, for example, under EC 1.3.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 6) or a trans-2-enoyi-CoA reductase classified, for example, under EC 1 ,3.1.38 or EC 1,3.1.8 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 7).

Pathways Usin NADH-Speeifie Emzym to Produce Nonadecanedioyl-C A Methyl Ester as ¾ Central Precursor Leading to C19 Building Blocks

In some embodiments, nonadecanedloyl-CoA methyl ester is synthesized from the central metabolite propanedioy!-CoA via eight cycles of methyl-ester shielded carbon chain elongation by conversion of propanedioyi-CoA to heptadecanedioyl-CoA methyl ester as described above; followed by conversion to 3-oxo-nonadecanedioyl~CoA methyl ester by a polypeptide having the activity of a β-ketoacyi-l ^' acp] synthase classified, for example, under EC 2.3. L- (e.g., EC 2.3.1 ,41 or EC 2.3,1.179) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 14-16) or a β-ketothiolase classified, for example, under EC 2.3.1.16 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 17); followed by conversion to 3- hydroxy-nonadecanedioyl-CoA methyl ester by a polypeptide having the activity of a 3~ hydroxyacyl-CoA dehydrogenase classified, for example, under EC 1.1.1 ,35 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 2); followed by conversion to 2,3- dehydrononadecanedioyl-CoA methyl ester by a polypeptide having the activity of an enoyl- CoA hydratase classified, for example, under EC 4.2.1.17 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 47); followed by conversion to nonadecanedioyl~CoA methyl ester by a polypeptide having the activity of a irans-2-enoyl-CoA reductase classified, for example, under EC 1.3.1 ,44 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 1 14 or SEQ ID NO: 115) or a polypeptide having the activity of a enoyl-facp] reductase classified, for example, under EC 1.3, 1.9 (e.g., e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ D NO: 46).

Pathways Using Noaadecaiiedioyl-CoA Methyl Ester or No¾adecaHedioyI~[acp] Methyl Ester as Central Precursors to Nonadecanedioic Acid

In some embodiments, nonadecanedioic acid is synthesized from the central precursor, nonadecanedioyl-[acp] methyl ester, by conversion of nonadecanedioyl-[aep] methyl ester to monomethyl nonadecanedioate by a polypeptide having the activity of a thioesierase classified, for example, under EC 3.1.1 ,2, EC 3,1.1.5, or EC 3.1.2.-, such as EC 3.1.2,14, EC 3.1 ,2.21, or EC 3.1 ,2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to nonadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 ,1 .1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonadecanedioic acid is synthesized from the central precursor, nonadeeanedioyhCoA methyl ester, by conversion of nonadecanedioyl-CoA methyl ester to monomeihyl nonadecanedioaie by a polypeptide having the activity of a thioesterase classified, for example, under EC 3.1.1.2, EC 3.1.1.5, or EC 3.1.2.-, such as EC 3.1.2.14, EC 3.1.2,21 , or EC 3.1 ,2.27, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%s sequence homology to the amino acid sequence of any one of SEQ ID NOs: 58-1 13 or SEQ ID NOs: 182-195; followed by conversion to nonadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51),

in some embodiments, nonadecanedioic acid is synthesized from the central precursor, nonadecanedioyl-CoA methyl ester, by conversion of nonadecanedioyl-CoA methyl ester to monomeihyl nonadecanedioaie by a olvpeptide having the activity of a CoA-transferase such as a glutaconate CoA-transferase classified, for example, under EC 2.8.3,12 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 40 or SEQ ID NO: 41) or a CoA-ligase classified, for example, under EC 6.2,1 .5; followed by conversion to nonadecanedioic acid by a polypeptide having the activity of esterase classified, for example, under EC 3.1.1.1 (e.g.. a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonadecanedioic acid is synthesized from the central precursor, nonadecanedioyl-CoA methyl ester, by conversion of nonadecanedioyl-CoA methyl ester to methyl 19-oxononadecanoate by a. polypeptide having the activity of an acetylaiing aldehyde dehydrogenase classified, for example, under EC 1.2,1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least. 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity o a succinate semialdehyde dehydrogenase classified, for example, under EC 1 ,2.1.76, or a polypeptide having the activity of an oxoglutaraie dehydrogenase classified, for example, under EC 1 ,2,1.52; followed by conversion to monomethyl nonadecanedioate by a polypeptide having the activity of a non-aeylating NAD- dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 11), a 7-oxohept noate dehydrogenase classified, for example, under EC 1.2.L- (e.g., a polypeptide having at least 5G%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6- oxohexanoate dehydrogenase classified, for example, under EC 1 ,2.1 ,- (e.g., a polypeptide having at least 50%, at. least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to nonadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, nonadecanedioic acid is synthesized from the central precursor, methyl 19-oxononadeeanoate, by conversion of methyl 19-oxononadecanoate to monomethyl nonadecanedioate by a polypeptide having the activity of a non- cylating NAD-dependent aldehyde dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 11), a 7- oxoheptanoaie dehydrogenase classified, for example, under EC 1.2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at. least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 12 or SEQ ID NO: 13), a 6-oxohexanoate dehydrogenase classified, for example, under EC 1,2.1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 9 or SEQ ID NO: 10), or an aldehyde dehydrogenase classified, for example, under EC 1.2.1.3 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 24); followed by conversion to nonadecanedioic acid by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

Pathways Using NoBa ecanedioyl-CoA Methyl Ester or Methyl 19-Oxon nadeeanoafe 'M a Central Precursor to 1 -Aminofi iiadecsE ate

In some embodiments, 19-aminononadecanoate is synthesized from the central precursor, nonadecanedioyl-CoA methyl ester, by conversion of nonadecanedioyl-CoA methyl ester to methyl 19-oxononadecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2.1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%) sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenas classified, for example, under EC 1.2.1.76. or a polypeptide having the activity of an oxoglutaraie dehydrogenase classified, for example, under EC 1.2.1 ,52; followed by conversion of methyl 19-oxononadecanoate to monomethyl 19-aminononadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2,6.1.1 L EC 2.6.1 ,13, EC 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ) ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 19-aminononadeeanoate to 19- aminononadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3,1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51), In some embodiments, 19-aminononadecanoate is synthesized from the central precursor, methyl 19-oxononadecanoate, by conversion of methyl 19-oxononadecanoate to monomethyl 1.9-amlnononadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6,1.-, such as EC 2,6.1.11. EC 2,6, 1.13. EC 2.6.1.18, EC 2.6.1.19, EC 2,6.1.29, EC 2,6.1.48, or EC 2,6.1.82, EC 4.1.1 ,64, or EC 5.4.3,8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 19-aminononadecanoate to 19- aminononadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO; 50 or SEQ ID NO: 51),

in some embodiments, 1 ~arninononadecanoate is synthesized from the central precursor, monomethyl nonadecanedioate, by conversion of monomethyl nonadecanedioate to methyl 19-oxononadecanoate by a polypeptide having the activity of a carhoxylaie reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70¾, or at least 85%o sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of methyl 19-oxononadecanoate to monomethyl 19-aminononadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6,1.11, EC 2.6.1.13, EC 2.6.1.18, EC 2,6.1.19, EC 2.6, 1.29, EC 2.6.1.48, or EC 2.6.1,82, EC 4.1 , 1.64, or EC 5,4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%> sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion of monomethyl 19- aminononadecanoate to 19-aminononadecanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1.1 , 1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51).

In some embodiments, 19~aminononadecanoate is synthesized from the central precursor, nonadecanedioic acid, by conversion of nonadecanedioic acid to 19- oxononadeeanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheim transferase classified, for example, under EC 2,7.8.-, such as 2.7,8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ Π) NOs: 54-57); followed by conversion of 19-oxononadecanoate to 19-aminononadecanoate by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1 ,-, such as EC 2.6.1.11, EC 2.6.1.13, EC 2.6,1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

Pathways Using 19~Aminonoiiadecaiioate, 19 ^» Hydr©xyiionadecaiioate, or 19-

Oxoffiosiadeeanoaie as Central Precursors to lNonadecaHe-i,19-DiaHiii e

in some embodiments, no.nadecane-l ,19-diamine is synthesized from the central precursor, 19-aminononadecanoate, by conversion of 19-aminononadecanoate to 19- aminononadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence horaology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopanteiheine transferase classified, for example, under EC 2.7.8.-, such as 2.7,8,7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 19-aminononadecanal to nonadecane-lj 9-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2,6.1.-, such as EC 2.6.1,11, EC 2.6.1.13, FX 2.6.1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181.

The carhoxylaie reductase encoded by the gene product of car and the phosphopantetheine transferase enhancer npt or sfp has broad substrate specificity, including terminal difunciionai C4 and C5 carboxyllc acids (Venkitasubrarnanian et /., Enzyme and Microbial Technology, 2008, 42, 130 - 137).

In some embodiments, nonadecane~l,1.9-dia ine is synthesized from the central precursor, 19-hydroxynonadecanoate, by conversion of 19~hydroxynonadeeanoate to 1.9- hydroxynonadecanal by a polypeptide having the activity of a c rboxylate reductase classified, for example, under EC 1,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 19-hydroxynonadecanal to 19-ammononadecano! by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.1 1, EC 2.6.1.13, EC 2.6.1.18, EC 2.6,1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%. or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16- 138 or SEQ ID NOs: 167-181 ; followed by conversion to 19-aminononadecanal by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1 .1, EC 1.1.1.2, EC 1.1.1.21, EC 1 ,1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23); .followed by conversion to nonadecane-l ,19~diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6, 1.-, such as EC 2.6.1.1 L EC 2,6, 1.13, EC 2.6.1.18, EC 2.6. L I 9, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181.

In some embodiments, nonadecane-I ,19-diamine is synthesized from the central precursor. 19-aminononadecanoate, by conversion of 19-aminononadecanoate to N19-acetyl- 19-aminononadecanoate by a polypeptide having the activity of an N~ cetyltransf erase such as a lysine N-acetyltransferase classified, for example, under EC 2.3.1 ,32 (e.g., a polypeptide having at least 50%, at least. 60%, at least. 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 53); followed by conversion to MI9-aeetyl~I9- aminononadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheim transferase classified, for example, under EC 2.7.8.-, such as 2.7.8.7 (e.g., a polypeptid having at least 50%, at least. 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to N19-aeetyl-l,19-diammononadecane by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1 ,1 1, EC 2,6.1.13, EC 2.6.1.18, EC 2.6, 1.19, EC 2.6.1.29, EC 2.6.1.48, or EC 2.6.1.82, EC 4.1.1.64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 1 16-138 or SEQ ID NOs: 167-181 ; followed by conversion to nonadecane~lJ.9~di amine by a polypeptide having the activity of an acelylp trescine deacylase classified, for example, under EC 3.5.1.62 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 42-45).

In some embodiments, nonadecane-1 ,19-di amine is synthesized from the central precursor, 19-oxononadecanoate, by conversion of 19-oxononadecanoate to 1,19- nonadecanedial by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2,7.8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57): followed by conversion to 19-aminononadecanal by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6. L-, such as EC 2,6.1.1 1, EC 2.6,1.13, EC 2.6.1.18, EC 2.6.1 ,19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6.1.82, EC 4.1.1 ,64, or EC 5.4.3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116-138 or SEQ ID NOs: 167-181 ; followed by conversion to nonadecane-1.19-diamine by a polypeptide having the activity of an aminotransferase classified, for example, under EC 2.6.1.-, such as EC 2.6.1.11, EC 2,6.1.13, EC 2.6, 1.18, EC 2.6.1.19, EC 2.6.1.29, EC 2.6.1 ,48, or EC 2.6.1 ,82, EC 4,1 ,1.64, or EC 5.4,3.8, or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85%s sequence homology to the amino acid sequence of any one of SEQ ID NOs: 116438 or SEQ ID NOs: 167-181.

Pathways Using oaadeeanedioic Acid or Noaadeeasedio l-CoA Methyl Ester as a Central Precurs r to 1 -Hydr xyKOisadeeaK aie

in some embodiments, 19-hydroxynonadecanoate is synthesized from the central precursor, nonadecanedioic acid, by conversion of nonadecanedioic acid to 19- oxononadecanoate by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99,6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8,-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion to 19-hydroxynonadecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1 ,1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1 ,1.1.258 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypenianoaie dehydrogenase classified, for example, under EC 1.1 .1.- (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21), or a polypeptid having the activity of a 4~hydroxyh tyrate dehydrogenase (e.g., a polypeptide having at least 50%o, at least 60%. at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23).

In some embodiments, 19-hydroxynonadecanoate is synthesized from the central precursor, nonadecanedioyl-CoA methyl ester, by conversion of nonadecanedioyl-CoA methyl ester to methyl 19-oxononadecanoate by a polypeptide having the activity of an acetylating aldehyde dehydrogenase classified, for example, EC 1.2, 1.10 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 18), a polypeptide having at least 50%, at least 60%, at least 70%>, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 19, a polypeptide having the activity of a succinate semialdehyde dehydrogenase classified, for example, under EC 1.2, 1.76, or a polypeptide having the activity of an oxoghiiarate dehydrogenase classified, for example, under EC 1.2.1 ,52; followed by conversion of methyl 19~oxononadeeanoate to 19~oxononadeeanoate by a polypeptide having the activity of an esterase classified, for example, under EC 3.1 , 1.1 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 50 or SEQ ID NO: 51); followed by conversion to 19-hydroxynonadecanoate by a polypeptide having the activity of a dehydrogenase classified, for example, under EC 1.1.1.- such as a 6-hydroxyhexanoate dehydrogenase classified, for example, under EC 1.1 ,1.258 (e.g., a polypeptide having at least 5Q%>, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 8), a polypeptide having the activity of a 5-hydroxypentanoate dehydrogenase classified, for example, under EC 1.1.1 .- (e.g., a polypeptide having at least 50%. at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 21 ), or a polypeptide having the activity of a 4-hydroxybutyrate dehydrogenase (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of SEQ ID NO: 23),

Pathways Using 19-HydroxyK nadeca»oaie as a Central Precursor to 1,19- onadeea&e iol

In some embodiments, 1,19-nonadecanediol is synthesized from the central precursor, 19-hydroxynonadeeanoate, by conversion of 19~hydroxynonadecanoate to 19- hydroxynonadeeanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1 ,2.99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a pkosphopantetheine transferase classified, for example, under EC 2.7,8.-, such as 2.7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 19-hydroxynonadecanal to 1 ,19-nonadecanediol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1 ,1.1.- (e.g.. EC 1.1 , 1.1, EC 1.1.1.2. EC 1.1.1 ,21 , EC 1.1 , 1.61, or EC 1.1.1.1 84) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 20-23).

Pathways Usisig i9~Aminoiionadeeasoate as a Central Precursor to 1 - Amiiiononadecanol

In some embodiments, 19-aminononadecanoi is synthesized from the central precursor, 19-aminononadeeanoate. by conversion of 19-aminononadecanoaie to 19- aminononadecanal by a polypeptide having the activity of a carboxylate reductase classified, for example, under EC 1.2,99.6 or a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 25-39 or SEQ ID NOs: 196-215 in combination with a polypeptide having the activity of a phosphopantetheine transferase classified, for example, under EC 2.7.8.-, such as 2,7.8.7 (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% sequence homology to the amino acid sequence of any one of SEQ ID NOs: 54-57); followed by conversion of 19-ammononadecanal to 19-ammononadecanol by a polypeptide having the activity of an alcohol dehydrogenase classified, for example, under EC 1.1.1.- (e.g., EC 1.1.1.1, EC 1.1.1.2, EC 1.1.1.21 , EC 1.1.1.61, or EC 1.1.1.184) (e.g., a polypeptide having at least 50%, at least 60%, at least 70%, or at least 85% .sequence homology to the amino acid sequence of any one of SEQ ID NOs; 20-23).

Cultivation Strategy

In some embodiments, the cultivation strategy entails achieving an aerobic, anaerobic, or micro-aerobic cultivation condition.

In some embodiments, the cultivation strategy entails nutrient limitation such as nitrogen, phosphate or oxygen limitation.

In some embodiments, a cell retention strategy using, for example, ceramic hollow fiber membranes can be employed to achieve and inaintain a high cell density during either fed-batch or continuous fermentation.

In some embodiments, the principal carbon source fed to the fermentation in the synthesis of one or more difunctional products having an odd number of carbon atoms (i.e., C _2n ÷3 building blocks, wherein n is an integer greater than or equal to one, such as C$ _t C ₇ , C _¾ Cn, Co, Ci5, C37, or Cj building blocks) can derive from biological or non-biological feedstocks,

In some embodiments, the biological feedstock can be, can include, or can derive from, monosaccharides, disaccharides. lignocelluiose, hemicellu!ose, cellulose, lignin, levulinic acid and formic acid, triglycerides, glycerol, fatty acids, agricultural waste, condensed distillers' solubles, or municipal waste.

In some embodiments, the feedstock is not glucose.

The efficient catabolism of crude glycerol stemming from the production of biodiesel has been demonstrated in several microorganisms such as Escherichia coli, Cupriavidus necator, Pseudomonas oleavorans, Pseudomonas putid and Yarrowia lipolytica (Lee et al , Appl Biochem. Biotechnoi, 2012, 166, 1801 - 1813; Yang et al.. Biotechnology for Biofoiels, 2012, 5: 13; Mdjnm et αί, Αρρί Microbiol. Biotechnoi., 2011, 90, 885 - 893). The efficient eatabolism of lignocelluiosic-derived levulinic acid has been demonstrated in several organisms such as Cupriavidus necator and Pseudomonas pittida in the synthesis of 3-hydroxyvaIerate via the precursor propanoyl-CoA (Jareniko and Yu, Journal of Biotechnology, 201 1, 155, 201 1, 293 -- 298; Martin and Prather, Journal of Biotechnology, 2009, 139, 61 - 67).

The efficient eatabolism of !ignin-derived aromatic compounds such as benzoate analogues has been demonstrated, in several microorganisms such as Pseudomonas putida, Cupriavidus necator (Bugg et al , Current Opinion in Biotechnology, 2011, 22, 394 - 400; Perez-Pantoja et al, FEMS Microbiol Rev., 2008, 32, 736 - 794).

The efficient utilization of agricultural waste, such as olive mill waste water has been demonstrated in several microorganisms, including Yorrowia Upolytica (Papanikolaou ei al, Bioresour. Tecknol , 2008, 99(7), 2419 - 2428),

The efficient utilization of fermentable sugars such as monosaccharides and disaccharides derived from eelMosic, hemicellulosic, cane and beet molasses, cassava, com, and other agricultural sources has been demonstrated for several microorganism such as Escherichia coli, Corynebacterium glutamicum and Lactobacillus delbrueckii and Laciococcus lactis (see, e.g., Hermann ei al, Journal of Biotechnology, 2003, 104, 155 - 172; Wee et al., Food TechnoL Bioiecknol, 2006, 44(2), 163 - 172; Ohashi et al , Journal of Bioscience and Bioengineering, 1999, 87(5). 647 - 654).

The efficient utilization of furfural, derived from a variety of agricultural lignocellulosic sources, has been demonstrated for Cupriavidus necator (Li et al , Biodegradation, 201 1, 22, 1215 - 1225).

in some embodiments, the non-biological feedstock can be, can include, or can derive from natural gas, syngas, CO ₂ H ₂ , methanol, ethanol, benzoic acid, non-volatile residue (NV ), a caustic wash waste stream from eyclohexane oxidation processes, or terephthalic acid / isophthalic acid mixture waste streams.

The efficient eatabolism of methanol has been demonstrated for the methylotrophic yeast Pichia pastoris (Yurimoto et al. , Int J Microbiol. , 201 1. 2011 : 101298).

The efficient eatabolism of ethanol has been demonstrated for Clostridium kluyveri (Seedorf et al. , Proc. Natl. Acad. ScL USA, 2008, 105(6) 2128 - 2133). The efficient catabolism of Ct¾ and H ₂ , which may be derived from natural gas and other chemical and petrochemical sources, has been demonstrated for Cupriavidus necator (Prybylski et al , Energy, Susiainability and Society, 2012, 2: 11),

The efficient catabolism of syngas has been demonstrated for numerous microorganisms, such as Clostridium !jungdahlii and Clostridium autoetkanogenum (Kopke et al, Applied and Environmental Microbiology, 2011, 77(15), 5467 - 5475).

The efficient catabolism of the non-volatile residue waste stream from eye! obex am; processes has been demonstrated for numerous microorganisms, such as Delftia aciaovorans and Cupriavidus necator (Ramsay et al, Applied and Environmental Microbiology, 1986, 52(1), 152 - 156).

In some embodiments, the host microorganism is a prokarvoie. For example, the prokaryote can be a bacterium from the genus Escherichia such as Escherichia coii; from the genus Clostridia _> such as Clostridium IjungdahlU, Clostridium autoeihanogermm, or Clostridium kl yverr, from the genus Corynebacteria _* such as Coryn bacterium glutamicum; from the genus Cupriavidus, such as Cupriavidus necator or Cupriavidus metalUdurans; from the genus Psetidomonas, such as Pseudomonas fluoresceins, Pseudomonas putida, or Pseudomonas oleavoram; from the genus Delftia such as Delftia acidovorans; from the genus Bacillus, such as Bacillus subtillis; from the genus Lactobacillus, such as Lactobacillus delhrueckii; or from the genus Lactococcus, such as Lactococcus lactis. Such prokaryotes also can be a source of genes to construct recombinant host cells described herein that are capable of producing one or more difunctionai products having an odd number of carbon atoms (i.e., C¾+3 building blocks, wherein « is an integer greater than or equal to one, such as Cj, C7, C$, C 1 ₁ , Co, Ci5, Cj ₇ , or Ci9 building blocks).

In some embodiments, the host microorganism is not Escherichia coll.

In some embodiments, the host microorganism is a eukaryote. For example, the eukaryote can be a filamentous fungus, e.g., a eukaryote from the genus Aspergillus, such as Aspergillus niger. Alternatively, the eukaryote can be a yeast, e.g., a eukaryote from the genus Saccharomyces such as Saccharomyces cerevisiae; from the genus Pichia such as Pichia pastoris; or from the genus Yarrowia such as Yarrowia lipolytica; from the genus Issatchenkia such as Issathenkia oriental-is; from the genus Debaryomyces such as Deharyomyces h msenii; from the genus Arxula such as Arxula adenoinivoram; or from the genus Kluyveromyces such as Kluyveromyces lactis. Such eukaryotes also can be a source of genes to construct recombinant host cells described herein that are capable of producing one or more difunctional products having an odd number of carbon atoms (i.e., C2 _H +3 building blocks, wherein n is an integer greater than or equal to one, such as C5, C _7j Cg, Cn, €53, C15, On, or Cj9 building blocks);

Metabolic Engineering

Metabolic pathway engineering has successfully been utilized by several groups to produce chemical commodities via fermentation processes. For example, recombinant strains expressing multiple exogenous genes and utilizing multi-step pathways not native to the strains have been developed. Recent advances in metabolic pathway engineering are summarized in, e.g., Chotani, Gopal, et al. "The commercial production of chemicals using pathway engineering," Biochimica et Biophysica Acta (BBA)-Protein Structure and Molecular Enzymology 1543,2 (2000): 434-455, Blombach. Bastian, and Bernhard J. Eikmanns. "Current knowledge on isobutanol production with Escherichia coli, Bacillus subtilis and Corynebaeterium glutamicum." Bioengineered Bugs 2.6 (2011): 346-350,, and Adkins, Jake, et al, "Engineering microbial chemical factories to produce renewable 'biomonomers'" Synthetic Biology Applications in Industrial Microbiology (2014): 31.

For example. Rathnasingh et al developed, a novel recombinant Escherichia coli SH254 strain that can produce 3-hydroxypropionic acid from glycerol via two consecutive enzymatic reactions. To develop the novel strains, Rathnasingh et al. inserted two plasmids, one encoding 5 exogenous genes utilized in. the enzymatic reactions, into an Escherichia coli SID 54 strain. See Rathnasingh, Chelladurai, et al. "Development and evaluation of efficient recombinant Escherichia coli strains for the production of 3-hydroxypropionic acid from glycerol" Bioiechnol Bioeng 104.4 (2009): 729-739.

in addition, Martin et al. engineered the expression of a synthetic amorpha~4,l l ~diene synthase gene and the nievalonate isoprenoid pathway from Sacckaromyces cerevisiae in Escherichia coli. See Martin, Vincent. XL et al "Engineering a nievalonate pathway in Escherichia coli for production of terpenoids," Nature Biotechnology 21.7 (2003 ): 796-802. The present document provides methods involving less than all the steps described for all the above pathways. Such methods can involve, for example, one, two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, or more of such steps. Where less than all the steps are included in such a method, the first, and in some embodiments the only, step can be any one of the steps listed.

Furthermore, recombinant hosts described herein can include any combination of the above enzymes such that one or more of the steps, e.g., one, two, three, four, five, six, seven, eight, nine, ten, or more of such steps, can be performed within a recombinant host. This document provides host cells of any of the genera and species listed and genetically engineered to express one or more (e.g., two, three, four, five, six, seven, eight, nine, ten, eleven, twelve, or more) recombinant forms of any of the enzymes recited in the document. Thus, for example, the host cells can contain exogenous nucleic acids encoding enzymes catalyzing one or more of the steps of any of the pathways described herein.

In addition, this document recognizes that where enzymes have been described as accepting CoA-activated substrates, analogous enzyme activities associated with [acp]-bound substrates exist that are not necessarily in the same enzyme class,

Also, this document recognizes that where enzymes have been described accepting (Rj-enantiomers of substrate, analogous enzyme activities associated with (S)-enantiomer substrates exist that are not necessarily in the same enzyme class.

This document also recognizes that where an enzyme is shown to accept a particular co-factor, such as NADPH, or a co-substrate, such as acetyl-CoA, many enzymes are promiscuous in terms of accepting a number of different co-factors or co-substrates in catalyzing a particular enzyme activity, Also, this document recognizes that where enzymes have high specificity for e.g., a particular co-factor such as NADH, an enzyme with similar or identical activity that has high specificity for the co-factor NADPH may be in a different enzyme class.

in some embodiments, the enzymes in the pathways outlined herein are the result of enzyme engineering via non-direct or rational enzyme design approaches with aims of improving activity, improving specificity, reducing feedback inhibition, reducing repression. improving enzyme solubility, changing stereo-specificity, or changing co-factor specificity. In some embodiments, the enzymes in the pathways outlined herein can be gene dosed (i.e., overexpressed by having a plurality of copies of the gene in the host organism), into the resulting genetically modified organism via episomal or chromosomal integration approaches.

In some embodiments, genome-scale system biology techniques such as Flux Balance Analysis can be utilized to devise genome scale attenuation or knockout strategies for directing carbon flux, to a difunctional product having an odd number of carbon atoms (i.e., a C _2N +3 building block, wherein n is an integer greater than or equal to one, such as a C5, C?, C9, Cii, C , Ci , On, or Ci9 building block).

Attenuation strategies include, but are not limited to, the use of transposons, homologous recombination (double cross-over approach), mutagenesis, enzyme inhibitors, and RNA interference (RNAi).

in some embodiments, fluxomic, metaho!omics, and transcriptomai data can be utilized to inform or support genome-scale system biology techniques, thereby devising genome scale attenuation or knockout strategies in directing carbon flux to a difunctional product having an odd number of carbon atoms (i.e., a C2n+3 building block, wherein n is an integer greater than or equal to one, such as a C-s, C _? , C9, CH, C13, CJS, Cn, or C19 building block).

In some embodiments, the host microorganism's tolerance to high concentrations of a difunctional product having an odd number of carbon atoms (i.e., a C _2N +3 building block, wherein n is an integer greater than or equal to one. such as a Cs, C?, C9, CH, C1 , Qs, Cn, or C19 building block) can be improved through continuous cultivation in a selective environment.

In some embodiments, the host microorganism's endogenous biochemical network can be attenuated or augmented to (1) ensure the intracellular availability of acetyl-CoA and propanedioyl-CoA, (2) create an NADH or NADPH imbalance that may only be balanced via the formation of one or more difunctional products having an odd number of carbon atoms (i.e., Ca _n+ 3 building blocks, wherein n is an integer greater than or equal to one, such as C5, C ₇ , C- , Cn, Co, Cjs, C17, or C Q building blocks), (3) prevent degradation of central metabolites, such as central precursors leading to and including one or more difunctional products having an odd number of carbon atoms (i.e., €-2»+ 3 building blocks, wherein n is an integer greater than or equal to one, such as C$, C ₇ , Cg, Cn, Co, CJS, Ci?, or C1 building blocks) and/or (4) ensure efficient efflux from the cell.

In some embodiments requiring intracellular availability of acetyl-CoA for €2,1+3 building block synthesis,, wherein n is an integer greater than or equal to one, endogenous enzymes catalyzing the hydrolysis acetyl-CoA such as short-chain length thioesterases can be attenuated in the host organism.

In some embodiments requiring condensation of acetyl-CoA and propanoyl-CoA for C-2n÷3 building block synthesis, wherein n is an integer greater than or equal to one, one or more endogenous β-ketothiolases catalyzing the condensation of only acetyl-CoA to acetoaeetyl-CoA, such as the endogenous gene products of atoB or ph A, can be attenuated. in some embodiments requiring the intracellular availability of aeetyhCo A for building block synthesis, wherein n is an integer greater than or equal to one, an endogenous phosphotransacetylase generating acetate such as pta can be attenuated (Shen et al, Appl Environ. Microbiol , 2011, 77(9):2905-2915).

In some embodiments requiring the intracellular availability of acetyl-CoA for C?.n+3 building block synthesis, wherein n is an integer greater than or equal to one, an endogenous gene in an acetate synthesis pathway encoding an acetate kinase, such as ack, can be attenuated.

In some embodiments requiring the intracellular availability of acetyl-CoA and NADH for C-2»+3 building block synthesis, wherein n is an integer greater than or equal to one, an endogenous gene encoding an enzyme that catalyzes the degradation of pyruvate to lactate, such as a lactate dehydrogenase encoded by IdhA, can be attenuated (Shen et al, 201 1, supra).

In some embodiments requiring the intracellular availability of acetyl-CoA and NADH for C -χ +3 building block synthesis, wherein n is an integer greater than or equal to one, endogenous genes encoding enzymes, such as a menaqiimol-fumarate oxidoreductase, that catalyze the degradation of phophoenolpyruvate to succinate, such as frdBC, can be attenuated (see, e.g., Shen et al,, 201 1 , supra).

In some embodiments requiring the intracellular availability of acetyl-CoA and NADH for C-2,,+3 building block synthesis, wherein n is an integer greater than, or equal to one, an endogenous gene encoding an enzyme that catalyzes the degradation of acetyi-CoA to ethanol, such as an alcohol dehydrogenase encoded by adhE, can be attenuated (Shen et al, , 201 1 , supra),

in some embodiments, where pathways require excess NADH co-factor for C¾n-;-3 building block synthesis, wherein n is an integer greater than or equal to one. a recombinant formate dehydrogenase gene can be overexpressed in the host organism (Shen et al , 201 1, supra). In some embodiments, said formate dehydrogenase is classified under EC 1.1 ,99,3 or EC 1.2.1 .2, such as the gene product of fdhF from, for example, Escherichia coli (see UniProtKB Accession No. P07658)

In some embodiments, where pathways require excess NADH or NADPH co-factor for Csii+s building block synthesis, wherein n is an integer greater than or equal to one, an endogenous transhydrogenase dissipating the co-factor imbalance can be attenuated.

In some embodiments, an endogenous gene encoding an enzyme that catalyzes the degradation of pyruvate to ethanol. such as a pyruvate decarboxylase, can be attenuated.

In some embodiments, an endogenous gene encoding an enzyme that catalyzes the generation of isobutanol, such as a 2-oxoacid decarboxylase, can be attenuated.

in some embodiments requiring the intracellular availability of acetyl-CoA for Can+s building block synthesis, wherein n is an integer greater than or equal to one, a recombinant acetyl-CoA synthetase, such as the gene product of acs from, for example, Escherichia coli (see UniProtKB Accession No, P27550 (SEQ ID NO: 151)) can be overexpressed in the microorganism (Satoh et al. , 1 Bioscience and Bioengineering, 2003, 95(4):335 - 341).

In some embodiments, carbon flux can be directed into the pentose phosphate, cycle to increase the supply of NADPH by attenuating an endogenous glucose-6-phosphate isomer ase, classified, for example, under EC 5,3.1.9.

In some embodiments, carbon flux can be redirected into the pentose phosphate cycle to increase the supply of NADPH by overexpression a 6-phosphogluconate dehydrogenase and/or a transketolase (Lee et al, 2003, Biotechnology Progress, 19(5), 1444 - 1449). In some embodiments, said 6-phosphogluconate dehydrogenase is classified, for example, under EC 1.1,1.44, such as the gene product of PGD from Homo sapiens (see UniProtKB Accession No. P52209 (SEQ ID NO: 152)). in some embodiments, said transketolase is classified, for example, under EC 2.2.1.1, such as the gene product of tktA from, for example, Escherichia coli (see UniProtKB Accession No. P27302 (SEQ ID NO: 153)).

In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a Cjn ₊ a building block, wherein n is an integer greater than or equal to one, a gene such as udhA encoding a puridine nucleotide transhydrogenase can be overexpressed in the host organisms (Brigham et al, Advanced Biofuels and Bioproducts, 2012, Chapter 39, 1065 - 1090).

In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a €2^-3 building block, wherein n is an integer greater than or equal to one, a recombinant glyceraldehyde-3-phosphate-dehydrogenase gene such as gapN can be overexpressed in the host organisms (Brigham et al, 2012, supra). In some embodiments, said glyceraldehyde-3-phospkate-dehydrogenase can be classified, for example, under EC 1.2.1.12, such as the gene product of gap A from, for example, Escherichia coli (see UniProtKB Accession No. P0A9B2 (SEQ ID NO: 155)).

In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a €¾ _η +3 building block, wherein n is an integer greater than or equal to one, a recombinant malic enzyme gene such as maeA or maeB can be overexpressed in the host organisms (Brigham et al , 2012). In some embodiments, said malic enzyme can be classified, for example, under EC LI .1 ,38, such as, for example, the gene product of maeA from, for example, Escherichia coli (see UniProtKB Accession No. P26616 (SEQ ID NO: 156)), or EC 1.1.1.40, such as, for example, the gene product of maeB from, for example, Escherichia coli (see UniProtKB Accession No. P76558 (SEQ ID NO: 157)).

In some embodiments, where pathways require excess NADPH co-factors in the synthesis of a C _¾+3 building block, wherein n is an integer greater than or equal to one, a recombinant glucose-6-phosphate dehydrogenase gene such as zwf cm be overexpressed in the host organisms (Lim et al , J, Bioscience and Bioengineering, 2002, 93(6), 543 - 549). In some embodiments, said glucose-6~phosphate dehydrogenase may be classified, for example, under EC 1.1.1.49, such as the gene product of zwf from, for example, Escherichia coli (see UniProtKB Accession No. P0AC53 (SEQ ID NO: 158)). In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a C2 _n +3 building block, wherein n is an integer greater than or equal to one. a recombinant fructose i, 6 diphosphatase gene such as jhp can be overexpressed in the host organisms (Becker et al , J. BiotechnoL, 2007, 132:99 - 109), In some embodiments, said fructose 1,6 diphosphatase may be classified, for example, under EC 3, 1.3.11, such as, for example, the gene product of ftp from, for example, Escherichia coli (see UniProt B Accession No. P0A993 (SEQ ID NO: 159).

In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a C n ₊₃ building block, wherein n is an integer greater than or equal to one, an endogenous t ose phosphate isomerase classified, for example, under EC 5,3.1.1 can be attenuated,

In some embodiments, where pathways require excess NADPH co-factor in the synthesis of a C2n+3 building block, wherein n is an integer greater than or equal to one, a recombinant glucose dehydrogenase such as the gene product of gdk can he overexpressed in the host organism (Satoh et al, J, Bioscience and Bioengineering, 2003, 95(4):335 - 341),

In some embodiments, endogenous enzymes facilitating the co ersion of ADPH to NADH can be attenuated, such as the NADH generation cycle that may be generated via inter-conversion of gluiamate dehydrogenases classified, for example, under EC 1.4,1.2 (NADH-specific) and EC 1.4,1.4 (NADPH-specific). For example, avoiding dissipation of an NADH imbalance towards Cs _n+ 3 building blocks, wherein n is an integer greater than or equal to one, a NADPH-specifie gh amate dehydrogenase can be attenuated.

In some embodiments, an endogenous glutamate dehydrogenase classified, for example, under EC 1.4,1.3 that utilizes both NADH and NADPH as co-factors can he attenuated.

In some embodiments, a membrane-bound enoyl-CoA reductase can be sohibilized via expression as a fusion protein to a small soluble protein such as a maltose binding protein (Gloerich et al, FEBS Letters, 2006, 580, 2092 - 2096).

In some embodiments using hosts that naturally accumulate polyhydroxyalkanoates, an endogenous polymer synthase enzymes can be attenuated in the host strain. In some embodiments, an L-alanine dehydrogenase, such as a Mycobacterium tuberculosis L-alanine dehydrogenase (see IJniProtKB Accession No. P9WQB1 (SEQ ID NO: 160)), can be overexpressed in the host to regenerate L-alanine from pyruvate as an amino donor for aminotransferase reactions,

In some embodiments, an L-glutamate dehydrogenase specific for the co-factor used to achieve co-factor imbalance can be overexpressed in the host to regenerate L-glutamate from 2-oxoglutarate as an amino dorsor for aminotransferase reactions. In some embodiments, said L-glutamate dehydrogenase is classified under, for example, EC 1.4.1.3, such as the gene product of GLUD from, for example, Homo sapiens (see UniProtKB Accession No, P00367 (SEQ ID NO: 161)). For example, to promote dissipation of the NADH imbalance towards C-2n÷3 building blocks, wherein n is an integer greater than or equal to one, a NADH-specific glutamate dehydrogenase can be overexpressed.

In some embodiments, enzymes such as a pimeloyl-CoA dehydrogenase classified, for example, under EC 1.3,1 .62; an acyl-CoA dehydrogenase classified, for example, under EC 13.8.7 or EC 1.3.8.1 ; and/or a glutary -CoA dehydrogenase classified, for example, under EC 1.3.8.6 that degrade central metabolites and central precursors leading to and including C s building blocks, wherein n is an integer greater than or equal to one, can be attenuated.

In some embodiments, endogenous enzymes activating C2n÷3 building blocks, wherein n is an integer greater than or equal to one, via Coenzyme A esterifi cation such as CoA- ligases (e.g., a pimeloyl-CoA synthetase) classified, for example, under EC 6.2.1.14 can be attenuated.

In some embodimen ts, a methanol dehydrogenase and a formaldehyde dehydrogenase can be overexpressed in the host to allow methanol cataboHsm via formate, in some embodiments, said methanol dehydrogenase may be classified, for example, under EC 1.1.1.244, such as the gene product of mdh from, for example, Bacillus methanolicus (see UniProtKB Accession No. P31005 (SEQ ID NO: 162)). In some embodiments, said formaldehyde dehydrogenase may be classified, for example, under EC 1.2.1.46, such as the gene product of fdhA from, for example. Biirkholderia pse domallei (see UniProtKB Accession No. A3P364 (SEQ ID NO: 163)). In some embodiments, an S-adenosylmethionine synthetase cars be overexpressed in the host to generate S~Adenosyl-L~methionine as a co-factor for S-adenosyl-L-methionine (SAM)-dependent methyitransferase. in some embodiments, said S-ade osylmethionine synthetase can be classified, for example, under EC 2.5.1 ,6, such as the gene product of metK from, for example, Escherichia coli (see UniProtKB Accession No. P0A817 (SEQ ID NO: 164)),

in some embodiments, the efflux of a (¾ _η +3 building block, wherein n is an integer greater than or equal to one, across the cell membrane to the extracellular media can be enhanced or amplified by genetically engineering structural modifications to the cell membrane or increasing any associated transporter activity for a C211+3 building block, wherein n is an integer greater than or equal to one.

The efflux of a diamine having an odd number of carbon atoms can be enhanced or amplified by overexpressing broad substrate range multidrug transporters, such as Bit from Bacillus subtilis (Woolridge et al., 1997, J. Biol, Chem., 272(14):8864 ^■■■ · 8866); AcrB and AcrD from Escherichia coli (Eikins & Nikaido, 2002, J. Bacterial., 184(23), 6490 - 6499), Nor A from Staphylococcus aereus (Ng et al,, 1994, Antimicrob Agents Chemother, 38(6), 1345 - 1355), or Bmr from Bacillus subtilis (Neyfakh, 1992, Antimicrob Agents Chemother, 36(2), 484 - 485), In some embodiments, the diamine having an odd number of carbon atoms may be pentane-l,5-diamme, heptane- 1 ,7-diamine, nonane-i,9-diamine, undecane-1 J 1- diamine, iridecane- 1,13 -diamine, pentadecane- 1,15 -diamine, heptadecane-l,17-diamine, or nonadecane- 1 , 19-diarnine,

The efflux of an aminocarboxvlate having an odd number of carbon atoms or a diamine having an odd number of carbon atoms can be enhanced or amplified by overexpressing the solute transporters such as the LysE transporter from Corynebacterium gl t mic m (Bellmann et al. , 2001, Microbiology, 147, 1765 - 1774), In some embodiments, the aminocarboxvlate having an odd number of carbon atoms may be 5-aminopentanoate, 7~ aminoheptanoate. 9-aminononanoate, l l-aminoundeeanoate, 13-arninotridecanoate, 15- aminopentadecanoate, 17-aminoheptadecanoate, or 19-aminononadecanoate. In some embodiments, the diamine having an odd number of carbon atoms may be pentane-1,5- diamine, heptane- 1 -diamine, nonane- 1 ,9-diamine, undecane- 1,1 1 -diamine, tridecane-1,13- dianiine, pentadeeane~l,15~diarnine, heptadecane-1 ,17-diamine, or nonadecane-l ,19-diarnine.

The efflux of a dicarboxylic acid having an odd number of carbon atoms can be enhanced or amplified by overexpressing a dicarboxylate transporter, such as the SucE transporter from Corynebactermm gl tarnicum (Hufm et aL , Appl. Microbiol. & Biotech, 89(2), 327 - 335). In some embodiments, the dicarboxylic acid having an odd number of carbon atoms is pentanedioic acid, b.eptanedioic acid, nonanedioic acid, undecanedioic acid, tridecanedioic acid, pentadecanedioie acid, heptadecanedioic acid, or nonadecanedioic acid,

Metabolicaily engineering recombinant hosts with various enzymes to produce final products has been successfully demonstrated by several groups. See, e.g., Blombach B et aL, Bioeng Bugs., 2011, 2(6):346-50 (teaching successful metabolic engineering of the last two steps of the Ehrlich pathway (by expression of genes encoding a broad range 2-ketoacid decarboxylase and an alcohol dehydrogenase) in recombinant hosts for the production of higher isobutanol); Adkins, J, et aL, Front Microbiol., 2012, 3:313 (summarizing numerous biomonomers (such as polyester building-blocks) that can be produced as a result of metabolic and pathway engineering in. various recombinant hosts); Chan, S. et aL, Bioresour TechnoL, 2012, 103(l):329-36 (teaching production of succinic acid from sucrose and sugarcane molasses by metabolicaily engineering E, co!i with sucrose-utilizing genes cscKB and cscA)); Lee, S, et at, Appl Biockem Biotechnol., 2012, 167(l):24-38 (teaching successful metabolic engineering of P. aeruginosa and E. coli for improving long-chain fatty acid production by co-expressing essential enzymes that are involved in the fatty acid synthesis metabolic pathway (accA and fabD) as well as fatty acyl-acyl carrier protein thioesterase gene); Rathnasingh, C. et al, Biotechnol Bioeng., 2009, 104(4):729-39 (teaching successful metabolic engineering of E. coli for producing 3-hydroxypropionic acid from glycerol by overexpression of glycerol dehydratas (DhaB) and aldehyde dehydrogenase (AldH) along with glycerol dehydratase reactivase (GDR)}.

Producing £211*3 Building Blocks Using a Recombinant Host

Typically, one or more difunctional products having an odd number of carbon atoms (i.e., C-2,,+3 building blocks, wherein n is an integer greater than or equal to one, such as C ₅₎ C7, C9. Cn, C33, C35, C37, or Cj9 building blocks) can be produced by providing a host microorganism and culturing the provided microorganism with a culture medium containing a suitable carbon source as described above, In general, the culture media and/or culture conditions can be such that the microorganisms grow to an adequate density and produce a difunctional product having an odd number of carbon, atoms (i.e., a ί¾π··3 building block, wherein n is an integer greater than or equal to one, such as a C5, C ₇ , C9, Cn, Ci ₃ , Cts,€57, or C39 building block) efficiently. For large-scale production processes, any method can be used, such, for example those described in Manual of Industrial Microbiology and Biotechnology, 2 ^nd Edition, Editors; A. L. Demain and J, E. Davies, ASM Press; and Principles of Fermentation Technology, P. F. Stanbury and A. Whitaker, Pergamon.

Briefly, a large tank (e.g., a 100 gallon, 200 gallon, 500 gallon, or more tank) containing an appropriate culture medium is inoculated with a particular microorganism. After inoculation, the microorganism is incubated to allow biomass to be produced. Once a desired biomass is reached, the broth containing the microorganisms can be transferred to a second tank, This second tank can be any size. For example, the second tank can be larger, smaller, or the same size as the first tank, Typically, the second tank is larger than the first such that additional culture medium can be added to the brot from the first tank. In addition, the culture medium within this second tank can be the same as, or different from, tha used in the first tank.

Once transferred, the microorganisms can be incubated to allow for the production of a difunctional product having an odd number of carbon atoms (i.e., a Om+3 building block, wherein n is an integer greater than or equal to one, such as a C ₅ , C ₇ , C¾ Cn,€53, C15, C17, or Ci9 building block). Once produced, any method can be used to isolate Can ₊ a building blocks. For example, Cjn ₊ n building blocks can be recovered selectively from the fermentation broth via adsorption processes. In the case of a dicarboxylic acid having an odd number of carbon atoms or an aminocarboxylate having an odd number of carbon atoms, the resulting eluate can be further concentrated via evaporation, crystallized via evaporative and/or cooling crystallization, and the crystals recovered via centrifugation. In the case of a diamine having an odd number of carbon atoms or a diol having an odd number of carbon atoms, distillation may be employed to achieve the desired product purity. In some embodiments,, a dicarboxylic acid having an odd number of carbon atoms may be pentanedioic acid, heptanedioie acid, nonanedioic acid, undeeanedioic acid, tridecanedioic acid, pentadecanedioic acid, hepiadecanedioic acid, or nonadeeanedioic acid.

In some embodiments, an aminocarboxyiate having an odd number of carbon atoms may be S-aminopertianoate. 7-aniinoheptanoate, 9-aminononanoate, 1 1 -amino undecanoate, 13 -amino tridecanoate, 15-aminopentadecanoate, 17-aminobeptadecanoate, or 19- an inononadccanoate,

In some embodiments, a diamine having an odd number of carbon atoms may be pentane- 1 ,5~diamine, heptane- 1 ,7-diamine, nonane- 1 ,9-dia ine, undecane- 1,11 -diamine, tridecane-L13-diamine, pentadecane~l,15~diamine, heptadeeane~l,17-diamine, or nonadecane- 1 _s 19-diamine,

In some embodiments, a dioi having an odd number of carbon atoms may be 1,5- pentanediol, 1,7-hepianediol, 1,9-nonanediol, 1,1 1 -undecanediol, 1,13-tridecanediol, 1,15- pentadecanediol, ί , 17-hepiadeeanediol, or 1,19-nonadecanedioL

Any of the recombinant hosts described herein may comprise a deletion in bioH. In some embodiments, the recombinant host does not express BioH. In some embodiments, the recombinant host may comprise a deletion in met J. In some embodiments, the recombinant host does not express MetJ.

The invention is further described in the following examples, which do not limit the scope of the invention described in the claims.

Enzyme Activity of Thioester ses Using Heptsmedioyl-f acp] er Hepianedioyl-[acp] Methyl Ester as a Substrate and Producing HoIo~ACP

The release of holo-ACP from pimeiyl~[acp] or pimelyl~[acp] methyl ester is catalyzed by the .following thioesterases:

The thioesterases were expressed and purified following procedures disclosed herein, The enzyme activity assay was performed in duplicate or triplicate in a buffer having heptanedioyi-Tacp] or heptanedioyl-[aep] methyl ester as substrate. The enzyme activity assay reaction was initiated by adding purified ihioesierase gene products to the assay buffer and incubating at 37 °C for 30 min. The release of holo-ACP was monitored by absorbance at 412 run. The absorbance associated with the empty vector control is very low, Many of the gene products of thioesterases accepted heptanedioyl-f aep] or heptanedioyl-[acp] methyl ester as substrate as confirmed via relative spectrophotometry (see FIG. 41), Notably, some thioesterases (e.g., 5ΤΈ, 6ΊΈ, 8ΊΈ, 1 ΠΈ, and 14 TE) synthesized holo-ACP more efficiently using heptanedioyl-f aep] methyl ester as substrate, when compared to using heptanedioyi- [acp] as substrate. See FIG, 41 ,

EXAMPLE 2

Enzyme Activity of Aminotransferase Using 7~OxoheptaB<Mte as a Substrate &ηύ Forming 7-Ammoheptaraoate

A sequence encoding an -terminal His-tag was added to the genes from Chromobacterium violacetm, Pseudomonas syringae, Rhodobacter sphaeroides, and Vibrio Fluvialis encoding the aminotransferases of SEQ ID NOs: 1 16 (GenBank Accession No. AAQ59697.1), 126 (GenBank Accession No. AAY39893.1), 127 (GenBank Accession No. ABA81135.1), and 128 (GenBank Accession No, AEA391.83.1), respectively such that N- terminal HIS tagged aminotransferases could be produced. Each of the resulting modified genes was cloned into a pET21a expression vector under control of the T7 promoter and each expression vector was transformed into a BL21 [DE3] E. coli host. The resulting recombinant E, coli strains were cultivated at 37°C in a 250mL shake flask culture containing 50 mL LB media and antibiotic selection pressure, with shaking at 230 rpm. Each culture was induced overnight at 16 °C using 1 mM IPTG.

The pellet from each induced shake flask culture was harvested via eentrifugation, Each pellet was resuspended and iysed via sonication. The cell debris was separated from the supernatant via eentrifugation and the cell free extract was used immediately in enzyme activity assays,

Enzyme activity assays in the reverse direction (i.e., 7-aminoheptanoate to 7- oxoheptanoate) were performed in a buffer composed of a final concentration of 50 mM HEPES buffer (pH = 7.5), 10 mM 7~animoheptanoate ₅ 10 mM pyruvate and 100 μΜ pyridoxyl 5' phosphate. Each enzyme activity assay reaction was initiated by adding cell free extract of the aminotransferase gene product or the empty vector control to the assay buffer containing the 7~aniinoheptanoate and incubated at 25°C for 4 h, with shaking at 250 rpm. The formation of L-alanine from pyruvate was quantified via RP-HPLC.

Each enzyme only control without 7-aminoheptanoate demonstrated low base line conversion of pyruvate to L-alanine. See FIG. 47. The gene product of SEQ ID NOs: 116

- ^*? 27 - (GenBank Accession No. AAQ59697.1), 126 (GenBank Accession No, AAY39893.1), 127 (GenBank Accession No. ABA81135.1), and 128 (GenBank Accession No, AEA39183.1) accepted 7-arainoheptanote as substrate as confirmed against the empty vector control See FIG, 48,

Enzyme activity in the forward direction (i.e., 7-oxoheptanoate to 7~aminoheptanoate) was confirmed for the aminotransferases of SEQ ID NO: 126 (GenBank Accession No. AAY39893.1), SEQ ID NO: 127 (GenBank Accession No, ABA81 135.1), and SEQ ID NO: 128 (GenBank Accession No. AEA391 S3,!). Enzyme activity assays were performed in a buffer composed of a final concentration of 50 mM HEPES buffer (pH = 7,5), 10 mM 7- oxoheptanoate, 10 mM L-alanine and 100 μΜ pyridoxyl 5' phosphate. Each enzyme activity assay reaction was initiated by adding a cell free extract of the aminotransferase gene product or the empty vector control to the assay buffer containing the 7-oxoheptanoate and incubated at 25°C for 4 h, with shaking at 250 rpm. The formation of pyruvate was quantified via RP- HPLC,

The gene product of SEQ ID NO: 126 (GenBank Accession No. AAY39893.1), SEQ ID NO: 127 (GenBank Accession No. ABAS 1135.1), and SEQ ID NO: 128 (GenBank Accession No. AEA39183.1) accepted 7-oxoheptanoate as a substrate as confirmed against the empty vector control. See FIG. 49, The reversibility of the aminotransferase activity was confirmed, demonstrating that the aminotransferases of SEQ ID NO: 126 (GenBank Accession No. AAY39893.1), SEQ ID NO: 127 (GenBank Accession No, ABAS 1135.1), and SEQ ID NO: 128 (GenBank Accession No. AEA39183.1) accepted 7-oxoheptanoate as substrate and synthesized 7-aminoheptanoate as a reaction product,

EXAMPLE 3

Enzyme Activity of Carboxylate Reductase Using Meptanedioate as a Substrate and Forming 7~Gxoheptaaoate

A sequence encoding a HIS-tag was added to the genes from Segniliparus rugosus and Segnillparus rotundus that encode the carboxylate reductases of SEQ ID NOs: 31 (GenBank Accession No. EFVl 1917.1) and 38 (GenBank Accession No. ADG98140.1), respectively, such that N~terminal HIS tagged carboxylate reductases could be produced. Each of the modified genes was cloned into a pET Duet expression vector along with a sfp gene encoding a HIS-tagged phosphopantetheine transferase from Bacillus subtilis, both under the T7 promoter. Each expression vector was transformed into a BL21[DE3] E, coll host and the resulting recombinant E. coli strains were cultivated at 37°C in a 250mL shake flask culture containing 50 mL LB media and antibiotic selection pressure, with shaking at 230 rpm. Each culture was induced overnight at 37 °C using an auto-induction media.

The pellet from each induced shake flask culture was harvested via centrifugation. Each pellet was resuspended and lysed via sonication, and the cell debris was separated from the supernatant via centrifugation. The carboxylate reductases and phosphopantetheine transferases were purified from the supernatant using Ni-affinity chromatography, diluted 10- fold into 50mM HEPES buffer (pH = 7,5), and concentrated via ultrafiltration.

Enzyme activity assays (i.e., from heptanedioate to 7-oxoheptan.oate) were performed in triplicate in a buffer composed of a final concentration of 50 mM HEPES buffer (pH = 7.5), 2 mM heptanedioate, 10 mM MgCl ₂ , 1 mM ATP, and 1 mM NADPH. Each enzyme activity assay reaction was initiated by adding purified carboxylat reductase and phosphopantetheine transferase gene products or the empty vector control to the assay buffer containing the heptanedioate and then incubated at room temperature for 20 min. The consumption of NADPH was monitored by absorbance at 340 nrn. Each enzyme only control without heptanedioate demonstrated low base line consumption of NADPH. See FIG. 42.

The gene products of SEQ ID NO: 31 (GenBank Accession No. EFV11917.1) and SEQ ID NO: 38 (GenBank Accession No, ADG9814G.1), enhanced by the gene product of sfp, accepted heptanedioate as substrate, as confirmed against the empty vector control (see FIG, 43), and synthesized 7-oxoheptanoate,

EXAMPLE 4

En yme Actsvsty of Carboxylate Reductase Using 7-Hydroxyheptanoafe as a Substrate asid Forming 7-Hy roxyfaeptanal

A sequence encoding a His-tag was added to the genes from Mycobacterium marinum, Mycobacterium smeg natis. Segniliparus rugosiis, Mycobacterium smegmatis, Mycobacterium massiliense, and Segniliparus rotundus that encode the carboxylate reductases of SEQ ID NOs: 25 (GenBank Accession No, ACC40567.1), 27 (GenBank Accession No, ABK75684.1), 29 (GenBank Accession No, AB .71854.1), 31 (GenBank Accession No. EFV11917.1), 37 (GenBank Accession No, EIV11143, 1), and 38 (GenBank Accession No. ADG9814Q.1) such that N-temiinal HIS tagged carboxylate reductases could be produced, Each of the modified genes was clotted into a pET Duet expression vector alongside a sfp gene encoding a His-tagged phosphopantetheine transferase from Bacillus subtilis, both under control of the T7 promoter.

Each expression vector was transformed into a BL21 [DE3] E. coli host and the resulting recombinant E. coli strains were cultivated at 37°C in a 25GmL shake flask culture containing 50 mL LB media and antibiotic selection pressure, with shaking at 230 rpm, Each culture was induced overnight at 37 °C using an auto-induction media.

The pellet from each induced shake flask culture was harvested via centrifugation. Kach pellet was resuspended and lysed via sonication. The cell debris was separated from the supernatant via centrifugation. The carbox late reductases and phosphopantetheine transferase were purified from the supernatant using Ni-affmity chromatography, diluted 10- fold into 50 mM HEPES buffer (pH = 7.5) and concentrated via ultrafiltration.

Enzyme activity (i.e., 7-hydroxyheptanoate to 7-hydroxyheptanal) assays were performed in triplicate in a buffer composed of a final concentration of 50 mM HEPES buffer (pH = 7.5), 2 mM 7-hydroxyheptanal, 10 mM MgCh, 1 n M ATP _; and 1 mM NADPH. Each enzyme activity assay reaction was initiated by adding purified carboxylate reductase and phosphopantetheine transferase or the empty vector control to the assay buffer containing the 7~hydroxyheptanoate and then incubated at room temperature for 20 min. The consumption of NADPH was monitored by absorbance at 340 nm. Each enzyme only control without 7- hydroxyheptanoate demonstrated low base line consumption of NADPH ^' . See FIG, 42,

The gene products of SEQ ID NOs: 25 (GenBank Accession No, ACC40567.1), 27

(GenBank Accession No. AB 75684.1), 29 (GenBank Accession No. AB 71854.1), 31 (GenBank Accession No, EFV11917.1), 37 (GenBank Accession No. EIV1 1143.1), and 38 (GenBank Accession No, ADG98140.1), enhanced by the gene product of sfp, accepted 7- hydroxyheptanoate as substrate as confirmed against the empty vector control (see FIG. 44), and synthesized 7-hydroxyheptanal. EXAMPLE 5

Enzyme Activity of Aminotransferase for T-Aminoheptaaol, Forming 7-OxoheptsMol

A nucleotide sequence encoding an N-terminal H s-tag was added to the

Chromobacteriwn violaceum, Pseudomortas syringae and Rkodobacier sphaeroides genes encoding the aminotransferases of SEQ ID NOs: 116 (GenBa k Accession No.

AAQ59697.1), 126 (GenBank Accession No. AAY39893.1), and 127 (GenBank Accession

No. ABA81 135,1), respectively such that N-terminal HIS tagged aminotransferases could be produced. The modified genes were cloned into a pET21 a expression vector under the T7 promoter. Each expression vector was transformed into a BL21 [DE3] E. coli host. Each resulting recombinant E. coli strain were cultivated at 37°C in a 250mL shake flask culture containing 50 mL LB media and antibiotic selection pressure, with shaking at 230 rpm. Each culture was induced overnight at 16°C using 1 mM IPTG.

The pellet from each induced shake flask culture was harvested via centrifugation.

Each pellet was resuspended and lysed via sonication. The cell debris was separated from the supernatant via centrifugation and the ceil free extract was used immediately in enzyme activity assays.

Enzyme activity assays in the reverse direction (i.e., 7-aminoheptanol to 7- oxoheptanol) were performed in a buffer composed of a final concentration of 50 mM HEPES buffer (pH ^::: 7.5), 10 mM 7-arninoheptanol, 10 mM pyruvate, and 100 uM pyridoxyl 5' phosphate. Each enzyme activity assay reaction was initiated by adding cell free extract of the aminotransferase gene product or the empty vector control to the assay buffer containing the 7-aminoheptanol and then incubated at 25 °C for 4 h, with shaking at 250 rpm. The formation of L-alanine was quantified via RP-HPLC.

Each enzyme only control without 7-aminoheptanol had low base line conversion of pyruvate to L-alanine. See FIG. 47.

The gene products of SEQ ID NOs: ί 16 (GenBank Accession No. AAQ59697.1), 126 (GenBank Accession No. AAY39893.1), and 127 (GenBank Accession No. ABA81135.1) accepted 7-aminoheptanol as a substrate as confirmed against the empty vector control (see FIG. 52) and synthesized 7-oxoheptanol as reaction product, Given the reversibility of the aminotransferase activity (see Example 2), it can be concluded that the gene products of SEQ ID NOs: 116 (GenBank Accession No. AAQ59697.1), 126 (GenBank Accession No. AAY39893.1), and 127 (GenBank Accession No, ABAS 1135,1) accept. 7-oxoheptanol as substrate and form 7~aminoheptanol.

EXAMPLE 6

£azyme Activity of Aminotransferase Using Heptane~l,7»diai»ine as a Substrate asid Forming 7-Amissohep†anaI

A sequence encoding an N-terminal His~tag was added to the Ckromohacterhim violaceum, Pseudomonas aeruginosa, Pseudomon s syringae, Rhodobacter sphaeroides, Escherichia coli, and Vibrio fl ialis genes encoding the aminotransferases of SEQ ID NOs: 116 (GenBank Accession No. AAQ59697.1), 1 19 (RefSeq Accession No. NP_417544.5), 125 (GenBank Accession No. AAG08191.1), 126 (GenBank Accession No. AAY39893.1), 127 (GenBank Accession No. ABA81135.1), and 128 (GenBank Accession No. AEA39183.1), respectively such that N-terminal HIS tagged aminotransferases could he produced. The modified genes were cloned into a pET21a expression vector under the T7 promoter. Each expression vector was transformed into a BL21 [DE3] E. coli host. Each resulting recombinant E. coli strain were cultivated at 37°C in a 250rnL shake flask culture containing 50 mL LB media and antibiotic selection pressure, with shaking at 230 rpm. Each culture was induced overnight at 16°C using 1 niM IPTG.

The pellet from each induced shake flask culture was harvested via centrifugation. Each pellet was resuspended and lysed via sonication. The cell debris was separated from the supernatant via centrifugation and the cell free extract was used immediately in enzyme activity assays.

Enzyme activity assays in the reverse direction (i.e., heptane- 1 ,7-diamine to 7- aminolieptanal) were performed in a buffer composed of a final concentration of 50 mM HEPES buffer (pH = 7.5), 10 niM heptane- 1 ,7-diamine, 10 mM pyruvate, and 100 μΜ pyridoxyi 5' phosphate, Each enzyme activity assay reaction was initiated by adding cell free extract, of the aminotransferase gene product or the empty vector control to the assay buffer containing the heptane- 1 ,7-diamine and then incubated at 25 °C for 4 h, with shaking at 250 rpm. The formation of L-alanine was quantified via RP-HPLC. Each enzyme only control without heptane- L7~diainine had low base line conversion of pyruvate to L-alanine. See FIG. 47.

The gene products of SEQ ID NOs: 116 (GenBank Accession No, AAQ59697.1), 119 (RefSeq Accession No. NP_417544.5), 125 (GenBank Accession No. AAG08191.1), 126 (GenBank Accession No. AAY39893.1), 127 (GenBank Accession No. ABA81 135.1), and 128 (GenBank Accession No. AEA39183.1) accepted heptane- 1 ,7~diamine as substrate as confirmed against the empty vector control (see FIG , 50) and synthesized 7-arninoheptanal as reaction product. Given the reversibility of the aminotransferase activity (see Example 2), it can be concluded that the gene products of SEQ ID NOs: 1 16. 1 19, and 125-128 accept 7- aminoheptanal as a substrate and form heptane- 1.7-diamme,

EXAMPLE 7

Enzyme Activity of Carhoxylate Reductase for N7-Acei !-7-AmiHohepianoaie _» Forming 7-Acetyl-7-AnsMoheptanaI

The activity of each of the N-terniinal His-tagged c rhoxylate reductases of SEQ ID NOs: 29 (GenBank Accession No. AB 71854.1), 37 (Genbank Accession No. EIV11143,1 ), and 38 (GenBank Accession No. ADG98140.1) (see Examples 4) for converting N7-acetyl-7- aminoheptanoate to N7~acetyl-7-aminoheptanal was assayed in triplicate in a buffer composed of a final concentration of 50 mM HEPES buffer (pH ^:::: 7.5), 2 mM N7~acetyl-7- ammoheptanoate, 10 mM MgCl _? ., I mM ATP, and 1 mM NADPH. The assays were initiated by adding purified carhoxylate reductase arid phosphopantetheme transferase or the empty vector control to the assay buffer containing the N7-acetyl-7-aminoheptanoate then incubated at room temperature for 20 min. The consumption of NADPH was monitored by absorbance at 340 nrn, Each enzyme only control without N7-acetyl-7-aminoheptanoate demonstrated low base line consumption of NADPH. See FIG, 42,

The gene products of SEQ ID NOs: 29, 37, and 38, enhanced by the gene product of sfp, accepted N7-acetyl-7-aminoheptanoate as substrate as confirmed against the empty vector control (see FIG. 45), and synthesized N7-acetyl-7-aminoheptanal. EXAMPLE 8

Enzyme Activity of Aminotransferase Using N7-Aeety!-l _s 7-Diaminoheptane, and Forming N7~AeetyI-7~Araim0heptaraal

The activity of the N-terminal His-tagged aminotransferases of SEQ ID NQs: 1 16 (GenBaiik Accession No. AAQ59697.1), 1 19 (RefSeq Accession No. NP_417544.5), 125 (GenBank Accession No. AAG08191.1), 126 (GenBank Accession No. AAY39893.1), 127 (GenBank Accession No. ABA81 135.1), and 128 (GenBank Accession No. AEA39183.1) (see Example 6) for converting N7-acetyl-l ,7-diaminoheptane to N7-acetyl-7-aminoheptanal was assayed using a buffer composed of a final concentration of 50 mM HEPES buffer ( H = 7.5), 10 mM N7-acetyl- 1 ,7-diaminoheptane, 10 mM pyruvate and 100 μΜ pyridoxyl 5' phosphate. Each enzyme activity assay reaction was initiated by adding a ceil free extract of the aminotransferases or the empty vector control to the assay buffer containing the N7- acetyl-1 ,7-diaminoheptane then incubated at 25 °C for 4 h, with shaking at 250 rpm. The formation of L~aianine was quantified via RP-HPLC.

Each enzyme only control without N7~acetyl~l, 7~diami.no heptane demonstrated low base line conversion of pyruvate to L-alanine. See FIG. 47.

The gene product of SEQ ID NOs: 116 (GenBank Accession No. AAQ59697.1), 119 (ReiSeq Accession No. NP _„ _417544.5), 125 (GenBank Accession No. AAG08191.1), 126 (GenBank Accession No. AAY39893.1), 127 (GenBank Accession No. ABA81135.1), and 128 (GenBank Accession No. AEA39183.1) accepted N7-acetyl- 1 ,7-diaminoheptane as a substrate as confirmed against the empty vector control (see FIG. 51) and synthesized N7- acetyl-7-aminoheptanai as reaction product.

Given the reversibility of the aminotransferase activity (see Example 2), the gene products of SEQ ID NOs: 116, 119, 125, 127, and 128 accept N7~acety!~7~ammoheptanal as substrate forming N7-acetyl- 1 ,7-diaminoheptane.

EXAMPLE 9

Enzyme Activity of Carboxylaie Reductase Using 7-Oxoheptanoate as & Substrate and

Forming Hepta&edial

The N-termiiial His-tagged carboxylaie reductase of SEQ ID NO: 38 (GenBank Accession No, ADG98140.1) (see Example 4) was assayed using 7-oxoheptanoate as substrate, The enzyme activity assay was performed in triplicate in a buffer composed of a final concentration of 50 raM HEPES buffer (pH ^::: 7,5), 2 mM 7-oxoheptarioate, 10 mM MgCl _¾ 1 mM ATP and 1 mM NADPH. The enzyme activity assay reaction was initiated by adding purified carboxylaie reductase and phospkopanteikeine transferase or the empty vector control to the assay buffer containing the 7-oxoheptanoate and then incubated at room temperature for 20 min, The consumption of NADPH was monitored by absorbance at 340 nm. The enzyme only control without 7-oxoheptanoate demonstrated low base line consumption of NADPH, See FIG. 42.

The gene product of SEQ ID NO: 38, enhanced by the gene product of sfp, accepted 7- oxoheptanoate as substrate as confirmed against the empty vector control (see FIG. 46) and synthesized heptanedial.

EXAMPLE 10

Enz me Activity of Esterase Using Monomethy! Hepianedioate as a Snbstrate and Forming Heptanediok Acid

A nucleotide sequence encoding a C-terminal His-tag may be added to the gene from. Bacillus s ht is encoding the esterase of SEQ ID NO: 50 (NCBI Reference Sequence: NP__388108.1) such that a C-terminal HIS tagged esterase is produced. The resulting modified gene is cloned into a pET28b+ expression vector under control of the T7 promoter and the expression vector is transformed into a BL21 [DE3] E. coli host. The resulting recombinant E. coli strain can be cultivated at 37°C in a 500mL shake flask culture containing 100 mL LB media and antibiotic selection pressure, with shaking at 230 rpm. Each culture is induced overnight at 18 °C using 0,3 mM iPTG.

The pellet from each induced shake flask culture can be harvested via centrifugation. Each pellet is resuspended and lysed via sonication. The cell debris is separated from the supernatant via centrifugation. The esterase can be purified from the supernatant using Ni~ affinity chromatography, buffer exchanged and concentrated into 20 mM HEPES buffer (pH ^:::: 7.5) via ultrafiltration and stored at 4 °C.

Enzyme activity assays converting monomethyl heptanedioate to heptanediok acid are performed in triplicate in a buffer composed of a final concentration of 25 mM Tris-HCl buffer (pH = 7.0) and 5 [mM] monomethyl heptanedioic acid. The enzyme activity assay reaction is initiated by adding esterases to a final concentration of 10 [μΜ] to the assay buffer containing the monomethyl heptanedioate and incubated at 30 ^o C for 1 h, with shaking at 250 φηι. The formation of heptanedioic acid can he quantified via LC-MS.

In this set. of esterase assays, a horse liver esterase (see, e.g., Craig et al, J, Am. Chem. Soc, 1958, 80 (7), 1.574-1579) and an esterase from Streptomyces diasiaiochromogenes (see, e.g., esiA from Streptomyces diastatochromogenes classified under EC 3.1.1 ,1 (UniProtKB access number Q59837), among other esterases, affected the methyl ester hydrolysis of a C compound.

EXAMPLE 11

Production of Heptanedioic Aeid in Geeetieally Modified Escherichi eoli (K12) Sirams

S-Adenosyl-methionine (SAM)-dependent methyltransferases (MTases) catalyse the transfer of methyl groups from SAM to a large variety of acceptor substrates ranging from small metabolites to bio-macromolecuies. See, e.g., Struck et a!, _s 2012, Chembiochem., 13(18):2642-55; see also FIGs. 1-3.

Many factors involved in. the SAM cycle may function to regulate the initial step of adding the methyl shield to propanedioyi-CoA or propanedioyl~[acp], which ultimately contribute to the altered production of heptanedioic acid. For example, MetK encodes for an enzyme that synthesizes SAM. Mtn encodes for S'-Methylthioadenosine nucleosidase, and mutants of Min accumulate 5 -deoxy adenosine and have impaired biotin synthase activity. On the other hand, tnetJ encodes a SAM co-repressor that represses the SAM biosynthetic enzymes, Me i represses expression of genes involved in methionine biosynthesis and is activated by increased levels of SAM. See, e.g., FIG. 53 for an illustration of representative factors involved in the SAM cycle. The Escherichia coli (KI2) strains carrying deletion in met J, and/or expression in metK and Mtn (see FIG. 54) were obtained following standard protocols to prepare genetically modified Escherichia coli (K12) strains. Strains with Acc were selected to allow comparison to no MetK/Mtn and AfadE backgfound. Acc was selected to avoid malonyl-CoA being limited as an increase of SAM was expected.

The mutant strains were tested for heptanedioic acid production in shake flask experiments (see FIGs. 55 and 56 for assay conditions and comments). The formation of heptanedioic acid was quantified via LC-MS. The strains with expression in metK and Mtn (INV0507) led to negligible increase in heptanedioic acid. Furthermore, deletion of met,/ coupled with expression of metK and Mtn (INV05Q8), did not lead to an increase in heptanedioic acid production. Unexpectedly, deletion in met J by itself (INV0509) increased the production of heptanedioic acid, Specifically, the data shows an increase in heptanedioic acid concentration from <1 mg/L (in the non-deletion strain) to around 3.5 mg/L (in the met J deletion strain). See FIG . 57, Addition of serine showed less heptanedioic acid despite better growth at 38 h.

Furthermore, HPLC was used to determine the S-adenosyi-L-methionine (SAM) and S-adenosyl-L-homocysteine (SAH) levels, which provides good separation and excellent detection limits and linear range (see, e.g., Wang et al, J Chromatogr B Biomed Sci Appl. 2001, 62(1):59~65; see also FIG. 58). Intracellular SAM was detected in all samples, and no SAH was detected by HPLC. For SAM levels, there is an increase from around 5-10 nmoles SAM/g wet cell weight (in the non-deletion strain) to 10-20 nmoles SAM/g wet cell weight (in the met J deletion strain), This depends on the timepoint hut the met/ deletion strain always produces more SAM, INV0508 supplemented with methionine resulted in the highest SAM levels (29.6 nmoles g WCW at t ^:::: 38h, 42% higher vs. no substrate; and 27% higher vs. serine feed), indicating that met J deletion coupled with expression of metK and Mtn boots SAM production. Furthermore, supplementation of serine/methionine increased SAM levels but there was no correlation to increase in heptanedioate yield. See FIGs. 57 and 59. EXAMPLE 12A

In Vitro and In Vivo Thioesterase Activity Screening

1,457 thioesterases were screened in a cell lysate in vitro assay for their ability to produce heptanedioate from heptanedioyl-ACP, In addition, 1,322 thioesterases were screened for their ability to produce monomethyl heptanedioate from heptanedioyl-ACP methyl ester in a cell lysate in vitro assay. Among the enzymes screened, seventy-six thioesterases (5.2% of enzymes screened) were identified as being able to carry out hydrolysis of heptanedioyl-ACP, and 147 thioesterases (11.1% of enzymes screened) were identified as being able to cany out the hydrolysis of heptanedioyl-ACP methyl ester.

The thioesterase library was received from Twist in IPTG-inducible T7~expression vectors and transformed into T7Express lysY/Iq. The resulting strains were induced with 0.4 niM IPTG, incubated overnight at 30 °C in 384- well plates, and lysed with Bugbuster containing Benzonase and Lysozyme. The cell lysates were separately screened for their ability to carry out the hydrolysis of heptanedioyl-ACP or heptanedioyl-[acp] methyl ester, Each thioesterase transformant strain was analyzed using biological duplicates (i.e., two colonies from each transformant), Activity was assessed by incubating the cell lysates in assay solution (50 mM tris pH 7.9, 100 raM NaCL 2% v/v glycerol, 500 μΜ DTNB, 0.33 mM aeyl-ACP substrate) in 384- well plates at 30 °C for 30 min. The absorbance of the wells was then measured at 410 nm, Data was analyzed by comparing to negative controls (gfp expression strain and vector-only) and a positive control (invista ΊΈ8 from Clostridium perfringens). Data were plotted in % activity compared to the negative control samples in that plate.

Thioesterases showing activity (judged as >10% "improvement" or % increase in Abs410 over the negative controls) were selected for further screening in vivo. A 175- member thioesterase library on IPTG-inducible T5~ l 5a expression vectors was co- transformed with BioC from Serratia marcescens (UniprotKB Accession No. P36571 (SEQ ID NO: 165)) expressed on a L-rhamnose inducible P(Rha)~pBR expression vector into both MG1655 rph+ AhioH and MG1655 rph+ AbioF. The plant thioesterase FatB2 from Cuphea hookeriana (UniprotKB Accession No. Q39514) was used as a positive control, FatB2 is a fatty acid acyl-ACP thioesterase with specificity toward Cg-Cio carbon chain lengths. Overexpression of FatB2 adversely affected cell growth, See FIG, 60 for plasmids.

The resulting strains were incubated in 0.5 mL of terrific broth (TB) supplemented with 1 mM IPTG and 2 mM L-rhamnose were added at the beginning of the experiment as inducers. The cultures were incubated overnight in 96-welS deep well plates at 37 °C in a shaking incubator. The cells were pelleted and culture supernatants were analyzed by LC-MS to determine the concentration of heptanedioate and monomethyl heptanedioate. Thioesterases active on heptanedioyl~ACP were detected in the AbioF strain via production of heptanedioate, and thioesterases active on methyl-heptanedioyl-ACP were detected via production of monomethyl heptanedioate in the kbioH strain, with impact determined as a percentage improvement over negative control.

Thioesterases for which no monomethyl heptanedioate production was detected scored as -100 improvement over negative control, Screening in a AhioH strain enabled approximately 14 mg/L of monomethyl heptanedioate production with FatB2 expression, about 10 mg/L higher than the negative control. Among the 146 thioesterases screened for impact on monomethyl heptanedioate production, four appeared to be promising and had a lower impact on cell growth than the positive control FatB2: UniProtKB Accession No. E4L0C9 (SEQ ID NO: 102); UniProtKB Accession No. A0A0B4Y4H4 (SEQ ID NO: 73); UniProtKB Accession No. F2JLT2 (SEQ ID NO: 74); and UniProtKB Accession No. A0A0B3WUQ1 (SEQ ID NO: 72). SEQ ID NO: 72 yielded an average of 48 mg/L monomethyl heptanedioate when the recombinant host expressing the thioesterase was grown in TB supplemented with 1 mM IPTG and 2 mM L-rhamnose, Further supplementation with serine and methionine appeared to inhibit monomethyl heptanedioate production.

EXAMPLE 12B

Additional Thioesterase Activity Screening

Thioesterase enzymes were selected using sequence similarity networks to sample a phylogenetically diverse set from the UniProtKB database and the ThYme database. From a selection of 1,956 enzymes, 1,457 thioesterases were screened on heptanedioy I.- ACP, with 76 hits, and 1,322 thioesterases were screened on heptanedioyl-ACP methyl ester, with 147 hits. Thioesierases corresponding to UniProtKB Accession Nos, A0A084JBW2 (SEQ ID NO: 88), D4YGM6 (SEQ ID NO: 95), R6RDZ9 (SEQ ID NO: 182), R6XLC3 (SEQ ID NO: 183), MIZIVO (SEQ ID NO: 184), C7ML86 (SEQ ID NO: 91), D0BKN0 (SEQ ID NO: 185), P44886 (SEQ ID NO: 186), and R5FQ35 (SEQ ID NO: 187) were notable for their activity toward heptanedioyl-ACP. Thioesierases corresponding to UniProtKB Accession Nos. A0A084JBW2 (SEQ ID NO: 88), B8I625 (SEQ ID NO: 89), G7V8P3 (SEQ ID NO: 76), C7ML86 (SEQ ID NO: 91), F5YIQ3 (SEQ ID NO: 92), A3DJY9 (SEQ ID NO: 75), H2FZ27 (SEQ ID NO: 93), POA8Z3 (SEQ ID NO: 60), A0A0D3V4E9 (SEQ ID NO: 94), and A4A3N9 (SEQ ID NO: 61 ) were notable for their activity toward heptanedioyl-ACP methyl ester. When the top 150 hits were subsequently screened in vivo, enzymes corresponding to UniprotKB Accession Nos. Q07792 (SEQ ID NO: 188), A0A0F7JXA5 (SEQ ID NO: 189), K5D7V3 (SEQ ID NO: 190), A0A0M9UHQ1 (SEQ ID NO: 191), A0A0F9W7B7 (SEQ ID NO: 86), A0A0C3EBX5 (SEQ ID NO: 192), A6D1N2 (SEQ ID NO: 82), A0A0B7DFD2 (SEQ ID NO: 193), A0A0B4Y4H4 (SEQ ID NO: 73), A4VL40 (SEQ ID NO: 194), E1WY53 (SEQ ID NO: 195), F7Z1I0 (SEQ ID NO: 108), and E1SPF5 (SEQ ID NO: 85) displayed considerable activity toward heptanedioyl-ACP, while enzymes corresponding to UniProtKB Accession Nos. E4L0C9 (SEQ ID NO: 102), AGA0B3WUQ1 (SEQ ID NO: 72), A3DJY9 (SEQ ID NO: 75), B1 ZXQ1 (SEQ ID NO: 78), A0A0B4Y4H4 (SEQ ID NO: 73), A0A0F7M706 (SEQ ID NO: 58), F2JLT2 (SEQ ID NO: 74), D5XAN2 (SEQ ID NO: 79), G7V8P3 (SEQ ID NO: 76), D6E2B1 (SEQ ID NO: 68), R6Q7V8 (SEQ ID NO: 80), D2TLW8 (SEQ ID NO: 81), and E1RAP4 (SEQ ID NO: 77) showed activity toward heptanedioyl-ACP methyl ester.

EXAMPLE 13

Selection of Aminotransferases with Specificity for Methyl 7~Oxohepta»oate Using Purified Enz mes

N-terminal His-tagged aminotransferases in T7 expression vectors were expressed in E. coli overnight at 30 °C for 16 hours. Cell pellets were resuspended in 50 mM HEPES pH 7.5, 100 mM NaCl, and 1 mM PLP. Lysis was performed using a microfluidizer and lysates were purified using Ni~NTA resin. A five-fold excess of pyruvate was added to the purified protein to convert any PMP present in the active site ΐο PLP. Pyruvate and imidazole were removed from purified protein by dialysis against 50 raM HEPES pH 7,5, 100 raM NaCl.

For titrations with 7-oxoheptanoate or methyl 7-oxoheptanoate. a coupled assay with lactate dehydrogenase was used to characterize activit using the pyruvate generated by the deamination of alanine, Reactions contained 100 m HEPES pH 7.5. 100 mM NaCl, 250 mM alanine, 0.8 mM NADU, 2.5 μ /ηι! lactate dehydrogenase, and 0.05 mg ml of aminotransferase. Concentrations of 7-oxoheptanoate and methyl 7-oxoheptanoate were varied until rate saturation or substrate inhibition was observed.

Similarly, for titrations with alanine, a coupled assay with lactate dehydrogenase was used to characterize activity using the pyruvate generated by the deamination of alanine. Reactions contained 100 mM HEPES pH 7.5, 100 mM NaCl, 2 mM 7-oxoheptanoate, 0.8 mM NADH, 2.5 }xg/ml lactate dehydrogenase, and 0,05 mg/ml of aminotransferase. Concentrations of alanine were varied from 0 mM to 200 mM.

In addition, to test g!utamate and aspartate as amine donors, a coupled assay with glutamate dehydrogenase was used to characterize activity with glutamate using the a- ketoglutarate generated by the deamination of glutamate. A coupled assay with malate dehydrogenase was used to characterize activity with aspartate using the oxaloacetate generated by the deamination of aspartate. Reactions contained 100 mM HEPES pH 7.5, 100 mM NaCl, 2 mM 7-oxoheptanoate, 100 mM aspartate or glutamate, 0,8 mM NADH, 0.4 mg/ml glutamate dehydrogenase or 0.15 mg/ml malate dehydrogenase, and 0.05 mg/ml of aminotransferase,

EXAMPLE 14A

in Vivo Aminotransferase Activity Screeni g

A 165-member aminotransferase library from Twist on IPTG-inducible T7- pUC expression vectors was co-transformed into T7 Express lysY/ ^' lq with a constitutive expression pJ23150-derived plasmid bearing the phosphopantelheinyl transferase sfp from Bacillus mbtilis, CAR__Srot from Invista and BioC from Serratla marcescens (UniprotKB Accession No. P36571 (SEQ ID NO: 165)). The resulting strains were incubated in 0.5 mi of rich media (terrific broth or LB containing 8% glycerol) supplemented with 0.4 mM IPTG and received 1 g/L monomethyl heptanedioate, 1 g/L heptanedioate, or no organic acid addition, depending on the experiment. Following incubation overnight at 37"C in a shaking incubator, cells were pelleted and the supernatants were analyzed using LC-MS for 7-aminoheptanoate or monomethyl 7-aminoheptanoate. A Vibrio fluvialis aminotransferase (UniProtKB Accession No. F2XBU9 (SEQ ID NO: 128)) was used as a positive control.

EXAMPLE 14B

Additional Aminotransferase Activity Screening

Aminotransferase enzymes were selected using sequence similarity networks to sample a phylogenetically diverse set from the UniProtKB database. Candidates were selected from each cluster, with 1 ,229 enzymes selected as being feasible for the desired reaction. Among these feasible enzymes, 167 were active toward 7-oxoheptanoate, Twelve of the top performing aminotransferases from in vitro analysis were taken forward for detailed kinetic analysis. Among these enzymes, aminotransferases corresponding to UniProtKB Accession Nos, A0A086YIZ0 (SEQ ID NO: 121), A0A01 1 UWB9 (SEQ ID NO: 131), A0A086MKC4 (SEQ ID NO: 133), A0A0E9ZHQ3 (SEQ ID NO: 123), A0A0H1A7R9 (SEQ ID NO: 134), B9L0N2 (SEQ ID NO: 117), H0I025 (SEQ ID NO: 124), J2TM48 (SEQ ID NO: 137), and Q7NWG4 (SEQ ID NO: 1 16) had a catalytic efficiency (K _cat /k _m ) favoring methyl 7- oxoheptanoate, whereas aminotransferases corresponding to UniProtK B Accession Nos. D7CVJ6 (SEQ ID NO: 130), A0A0HIAH98 (SEQ ID NO: 122), D7A1Z2 (SEQ ID NO: 135), G7Z3P2 (SEQ ID NO: 136), and K.2KXB1 (SEQ ID NO: 138) had a preference for 7- oxoheptanoate. In parallel, 120 aminotransferases were also screened in vivo for the conversion of 7-oxoheptanoate to 7~aminohepta.noa.ie, and thirty of these generated more 7- aminoheptanoate than the positive control (UniProtKB Accession No, F2XBU9 (SEQ ID NO: 128)), Examples of aminotransferases with higher 7-aminoheptanoate yields include enzymes corresponding to UniProtKB Accession Nos. A0A0E9ZHQ3 (SEQ ID NO: 123), B6ISI5 (SEQ ID NO: 168), A0A0F9UFF8 (SEQ ID NO: 169), C7LZG4 (SEQ ID NO: 170), Ι3ΊΉ77 (SEQ ID NO: 171), V7D492 (SEQ ID NO: 172), A0A0C6G014 (SEQ ID NO: 173), G3BAK1 (SEQ ID NO: 174), A0A081B6K8 (SEQ ID NO: 175), R5HDC3 (SEQ ID NO: 176), A3U3W9 (SEQ ID NO: 177), A0A086YIZ0 (SEQ ID NO: 121), K2KXB1 (SEQ ID NO: 138), B9L0N2 (SEQ ID NO: 117), A0A0H1AH98 (SEQ ID NO: 122), A0A059IS31 (SEQ ID NO: 178), D7VKX2 (SEQ ID NO: 179), F2XBU9 (SEQ ID NO: 128), F5Y1J0 (SEQ ID NO: 180), and A0A061M4Q7 (SEQ ID NO: 181).

N-tetminal, His-tagged carboxylate reductases were co-expressed with a phosphopanteiheine transferase (sfp) in T7 expression vectors in E.coli overnight at 28 C for 16 hours, The cell pellets were re~suspended in 50 mM potassium phosphate pH 7.5, 300 mM NaCl, 10% (w/V) glycerol. Lysis was performed using a microfluidizer (two passes at 10,000 PSI), The lysates were clarified by centrifugation (12,000 x g for 1 hour) and purified using fast protein liquid chromatography (FPLC) with Ni~NTA resin employing a gradient of 0 to 500 mM imidazole. Peak fractions were analyzed for activity, and purity was confirmed via SDS-PAGE (95% pure).

For titrations with heptanedioate/monomethyl heptanedioate, 7-oxoheptanoate/methyl 7-oxohepianoate, and 7-aminoheptanoate/monomethyl 7~aminoheptanoate, the reaction mixtures contained 100 mM Tris pH 7.5, 1 mM EDTA, 2 mM β-mercaptoethanol, 1 mM MgCla, 10% (w/v) Glycerol, 0,5 mM NADPH, 1 mM ATP, and 5 of carboxylate reductase (co-expressed with phosphopanteiheine transferase). The reactions were monitored by following the decrease in absorbance at 340 nm. Concentrations of the substrates were varied until rate saturation was observed. For methyl 7-oxoheptanoate, 7-aminoheptanoate, and monomethyl-7~aminohepianoate, the substrates were titrated up to 20 mM without detecting any activity.

For reactions to test degree of phosphopantetheinylation, carboxylate reductases and carboxylate reductases co-expressed with phosphopanteiheine transferase were incubated with phosphopanteiheine transferase and 1 mM acetyl-CoA for 1 hour at 37 C. The activity of the enzymes were then determined by adding 5 μΕ of each mixture to 95 \xL of 100 mM Tris pH 7.5, 1 mM EDTA, 2 mM β-mercaptoethanol, 1 mM MgCb, 10% (w/v) Glycerol, 0,5 mM NADPH, I mM ATP, and 5 mM monomethyl heptanedioate. The reactions were monitored by following the decrease in absorbance at 340 nm. To screen carhoxylate reductases in vitro, a 570-member carboxylase reductase library from Twist on IPTG-inducible T7~ pUC expression vectors was co-transformed into T7 Express lysY/Iq with the phosphopantetheinyl transferase sfp from Bacillus subtilis on a T7-pCDF expression vector. The Invista CAR Srug and Srug69 were used as positive controls. Biological duplicates of the resulting strains were incubated in 0,5 ml of terrific broth supplemented with 0.4 mM IPTG in 96-well deep well plates. The plates were incubated overnight at 30 °C in a shaking incubator. 150 μΐ of the cultures were transferred to new 96-well deep well plates to which 150 μΕ BugBuster supplemented with lysozyme (l .uL/mL), benzonase (Ι Ι,/mL), and PMSF (0.5 mM) was added and incubated for 10 min at room temperature. The cell debris was pelleted and 10 Τ of the supernatants was assayed for activity.

In addition, NADPH oxidation assays were performed on the supernantants in 96-well plates with 100 μΐ reaction volumes. The assay mixture contained: 50 mM HEPES pH 7,5, 10 mM MgCl _2; 10 % glycerol, 1 mM ATP, 0.2 mM NADPH, 5 mM monomethyl heptanedioate or heptanedioate. The assay was performed at 37 °C and readings at 340 nm were taken at 30, 120, and 180 minutes.

To screen carhoxylate reductases in vivo, a 497-member carhoxylate reductase library from Twist on IPTG-inducible T7- pUC expression vectors was co-transformed into T7 Express lysY Iq with the phosphopantetheinyl transferase sfp from Bacillus subtilis on a T7- pCDF expression vector. Biological duplicates of the resulting strains were incubated in 0,5 ml, of terrific broth containing either 1 g/L heptanedioate or 1 g/L monomethyl heptanedioate and supplemented with 0.4 mM IPTG in 96-well deep well plates. The plates were incubated for 30 h at 30 °C in a shaking incubator. The cells were then pelleted, and culture supernatants from the same strains in media containing either heptanedioate or monomethyl heptanedioate. were combined. The combined supematants were analyzed by LC-MS to determine the concentration of heptanedioate and monomethyl heptanedioate. Carhoxylate reductase activity on heptanedioate or monomethyl heptanedioate was reflected by consumption of the respective substrate in the medium. EXAMPLE 16B

Additional Carboxylase Reductase Activity Screening Carboxylate reductase enzymes were selected using sequence similarity networks to sample a phyiogeneticaSly diverse set from the UniProtKB database. Selecting candidates from each cluster, a total of 541 enzymes were screened in vivo to convert monomethyl heptanedioate and heptanedioic acid. Enzymes corresponding to UniProtKB Accession Nos. A0A0G4ID64 (SEQ ID NO: 196), A0A0H5CAG1 (SEQ ID NO: 197), W3XHR4 (SEQ ID NO: 198), Q1 8660 (SEQ ID NO: 199), I7MB41 (SEQ ID NO: 200), A0A087SHC7 (SEQ ID NO: 201), A0A068SDQ8 (SEQ ID NO: 202), A0A0K1PNT5 (SEQ ID NO: 203), A0A0J9XGX9 (SEQ ID NO: 204), W6MHS7 (SEQ ID NO: 205), and G4YTV4 (SEQ ID NO: 206) preferred acting upon heptanedioic acid to form 7-oxoheptanoate; many of these enzymes displayed little activity for monomethyl heptanedioate. However, the tested enzymes with the most monomethyl heptanedioate activity included enzymes corresponding to UniProtKB Accession Nos. Q1ZXQ4 (SEQ ID NO: 207), F1KXI1 (SEQ ID NO: 208), A0A034UK40 (SEQ ID NO: 209), A0A0C7BIS0 (SEQ ID NO: 210), Q4N8F1 (SEQ ID NO: 21 1), A0A0M3J210 (SEQ ID NO: 212), T1 EG09 (SEQ ID NO: 213), A0A0F4ES51 (SEQ ID NO: 214), and A0A0H2RRC5 (SEQ ID NO: 215).

EXAMPLE 17

Alcohol Dehydrogenase Activity Seree&iisg

Sixty-eight potential alcohol dehydrogenase genes from E. co!i were cloned into

I TG -inducible T5-expression vectors. The enzyme library was transformed into MG1655 rph+ AybbO &yahK Aahr &adhP AyqhD AyiaY. Biological triplicates of strains were grown in 0.5 mL terrific broth supplemented with 0.5mM IPTG for induction. The culture incubation was performed in 96-we!l deep well plates for 4 h at 37 °C in a shaking incubator. The cells were pelleted by centrifugalion at 4 °C, and the supernatant was removed, The cell pellets were resuspended in 200 μΕ BugBuster supplemented with, lysozyme (l .L/mL), benzonase (I pL/niL), and PMSF (0.5 mM) and incubated for 10-20 min at room temperature, after which the cell debris was pelleted, leaving the supernatant as the crude extract. Assays were performed on the supematants in 96-well plates in 200 pL reactions, The assay mixture contained 50 mM MOPS, 0.2mM NAD(H) or NADPH, and 10 μΐ, crude extract. Reactions were initiated by the addition of 2 mM 7-oxoheptanoate, Prior to initiation of the assay, the enzymes were equilibrated at 37 °C for 3 min. After initiation, the decrease or increase in absorbance monitored at 340 nm was measured continuously for 3 min. Each plate was measured again after 30 min has passed.

OTHER EMBODIMENTS

It is to he understood that, while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims.